AU710881B2 - 1-methylcarbapenem derivatives - Google Patents

Abstract

A 1-methylcarbapenem compound represented by the following formula: <CHEM> Äwherein R<1> represents a hydrogen atom or a lower alkyl group, R<2> represents a hydrogen atom or a lower alkyl group, R<3> represents a hydrogen atom, a lower alkyl group, a lower alkyl group which has a substituent, a cycloalkyl group or a group of formula -C(=NH)R<4> (in which R<4> represents a hydrogen atom, a lower alkyl group or an amino group)Ü; or a pharmacologically acceptable salt or derivative thereof. ÄAdvantagesÜ The 1-methylcarbapenem compounds of the present invention exhibit excellent antibacterial activity and are therefore effective as a preventive or remedy of infections.

Description

P:\OPER\PDB\24076-97.286 6/8/99 clinical situation, however, resistant strains against it in Pseudomonas aeruginosa and the like have started to be recognized.

There is accordingly an increasing need for agents which exhibit stronger and well-balanced antibacterial activity against a wide range of bacteria, including strains of Pseudomonas aeruginosa which exhibit resistance against meropenem, and are free from nephrotoxicity. In Japanese Patent Application Kokai No. Hei 5-310740, Japanese Patent Application Kokai No. Hei 5-339269, Japanese Patent Application Kokai No. Hei 6-172356 and Japanese Patent Application Kokai No. Hei 6-199860, 1-methylcarbapenem derivatives synthesized for satisfying the above-described need are disclosed.

With a view toward overcoming the above-described defects of 1methylcarbapenem derivatives, the present inventors carried out investigations. As a result, it has been found that compared with the conventional 1-methylcarbapenem derivatives, compounds of the present invention have superior antibacterial activity, are more stable against dehydropeptidase I, have improved recovery rates in urine and are superior in pharmacokinetics such as half-life in blood. It has also been found that the compounds of the present invention have low nephrotoxicity and are therefore effective as an antibacterial agent used for the treatment of prevention (particularly treatment) of bacterial infections.

In Japanese Patent Application Kokai No. Hei 5-310740, Japanese Patent Application Kokai No. Hei 5-339269 and Japanese Patent Application Kokai No.

Hei 6-172356, 3 -(aminomethyl)pyrrolidine-containing compounds [a compound represented by the formula wherein

R

2 and R 3 are hydrogen atoms at the same time, and the like] are disclosed, by they do not contain any disclosure about the preparation of compounds which belong to the present invention and have a substituent at the amino part of the 3 -(aminomethyl)pyrrolidine group.

The present invention provides a 1-methylcarbapenem compound R,5S,6S)-6-[(1R)-1-hydroxyethyl]-l-methyl-2-[(2S,4S)-2-[(3R)-3methylaminomethylpyrrolidin-1l-ylcarbonyl]pyrolidin-4-ylthio]- -carbapen-2-em-3carboxylic acid having excellent antibacterial activity, pharmacologically acceptable salts or derivatives thereof; compositions for the prevention or treatment of bacterial infections which P:\OPER\PDB\24076-97.286 6/8/99 comprises said compound, salts and derivatives as an effective ingredient; use of said compound, salts or derivatives for the preparation of a medicament used for the prevention or treatment of bacterial infections; a method for the prevention or treatment of bacterial infections which comprises administering a pharmacologically effective amount of said compound, salts and derivatives to warm-blooded animals; and processes for the preparation of said compound, salts or derivatives.

The 1-methylcarbapenem derivatives of the present invention are represented by formula: 4 OH6 CH 3 2 CON 3 R2 CH3 N4 2 R3 N R1

COOH

(I)

wherein: R' represents a hydrogen atom or a lower alkyl group;

R

2 represents a hydrogen atom or a lower alkyl group; and

R

3 represents a hydrogen atom, a lower alkyl group, a lower alkyl group which has 1 to 3 substituents (each of said substituents is a hydroxyl group, a halogen atom, a carbamoyl group, a carbamoyl group substituted by 1 or 2 lower alkyl groups, a carbamoyloxy group, a carbamoyloxy group substituted by 1 or 2 lower alkyl groups, a lower alkoxy group, an amino group or an amino group substituted by 1 or 2 lower alkyl groups), a cycloalkyl group or a group of formula

NH)R

4 (in which R 4 represents a hydrogen atom, a lower alkyl group or an amino group), or

R

2 and R 3 taken together represent an alkylene group which is optionally interrupted by one oxygen, nitrogen or sulfur atom (said nitrogen atom may be substituted by a lower alkyl group); with the proviso that R 2 and R 3 do not represent hydrogen atoms at the s AL same time.

In the above description, the "lower alkyl group" in the definitions ofR 1

R

2

R

3 and R 4 is a straight or branched

C

1 -4 alkyl group. Examples of such a group include methyl, ethyl, n-propyl, isopropyl, n-butyl and t-butyl groups, of which a methyl or ethyl group is preferred, a methyl group being more preferred.

Examples of the "halogen atom" in the definition of R 3 include fluorine, chlorine and bromine atoms, of which a fluorine atom is preferred.

Examples of the "carbamoyl group substituted by 1 or 2 lower alkyl groups" in the definition ofR 3 include methylcarbamoyl, ethylcarbamoyl, dimethylcarbamoyl and diethylcarbamoyl groups, of which a methylcarbamoyl or dimethylcarbamoyl group is preferred.

Examples of the "carbamoyloxy group substituted by 1 or 2 lower alkyl groups" in the definition ofR 3 include methylcarbamoyloxy, ethylcarbamoyloxy, dimethylcarbamoyloxy and diethylcarbamoyloxy groups, of which a methylcarbamoyloxy or dimethylcarbamoyloxy group is preferred.

Examples of the "amino group substituted by 1 or 2 lower alkyl groups" in the definition ofR 3 include methylamino, ethylamino, dimethylamino and diethylamino groups, of which a methylamino or dimethylamino group is preferred.

The "lower alkoxy group" in the definition ofR 3 is a C 1 4 alkoxy group and examples include methoxy, ethoxy, propoxy and butoxy groups, of which a methoxy group is preferred.

Preferred examples of the "substituent" of the "lower alkyl group having 1 to 3 substituents" in the definition ofR 3 include a hydroxy group, a fluorine atom, a carbamoyl, methylcarbamoyl, dimethylcarbamoyl, carbamoyloxy, methoxy, amino, methylamino or dimethylamino group.

Examples of the above-described "lower alkyl group having 1 to 3 substituents" include 2 -hydroxyethyl, 3-hydroxypropyl, 2 -hydroxypropyl, 2,3-dihydroxypropyl, fluoromethyl, difluoromethyl, trifluoromethyl, 2-fluoroethyl, 2 2 2 -trifluoroethyl, chloromethyl, dichloromethyl, trichloromethyl, 2-chloroethyl, carbamoylmethyl,

N-

methylcarbamoylmethyl, NN-dimethylcarbamoylmethyl, 2-carbamoylethyl, 2-(N- 97162p2.doc

P

7 9 56 9 /FP-9716(PC'I)/tsaig/English translation/07.10.98 methylcarbamoyl)ethyl, 2 -(NN-dimethylcarbamoyl)ethyl, 2 -carbamoyloxyethyl, 2- (N,N-dimethylcarbamoyloxy)ethyl, 2 -methoxyethyl, 2-ethoxyethyl, 2-aminoethyl, 3aminopropyl, 2-aminopropyl, 2 -(methylamino)ethyl, 3-(methylamino)propyl 2- (dimethylamino)ethyl and 3 -(dimethylamino)propyl groups; of which a 2 -hydroxyethyl, 2-fluoroethyl, 2 2 2 -trifluoroethyl, carbamoylmethyl, 2 -carbamoylethyl,

N-

methylcarbamoylmethyl, NN-dimethylcarbamoylmethyl, 2 -methoxyethyl, 2carbamoyloxyethyl, 2 -aminoethyl, 2 -(methylamino)ethyl or 2 -(dimethylamino)ethyl group is preferred, a 2 -hydroxyethyl, carbamoylmethyl, 2-aminoethyl or 2- (methylamino)ethyl group being more preferred.

The "cycloalkyl group" in the definition of R 3 is a C 3 6 cycloalkyl group and examples of such a group includes cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl groups; of which a cyclopropyl or cyclobutyl group is preferred, a cyclopropyl group being more preferred.

Examples of the "group represented by the formula -C(=NH)R 4 include formimidoyl, acetimidoyl, propioimidoyl and amidino groups; of which a formimidoyl, acetimidoyl or amidino group is preferred.

The "alkylene group" of the "alkylene group which is optionally interrupted by one oxygen, nitrogen or sulfur atom" in the definition of the groups of R 2 and R 3 taken together is a straight or branched C2-6 alkylene group and examples of such a group include ethylene, propylene, trimethylene, 1 -methyltrimethylene, 2 -methyltrimethylene, tetramethylene, I -methyltetramethylene, 2 -methyltetramethylene and pentamethylene groups.

Examples of the alkylene group which is interrupted by one oxygen, nitrogen or sulfur atom include ethyleneoxyethylene(CH 2

CC

H

CH

2 ethyleneaminoethylene (CH2CH 2

NHCH

2

CH

2 ethyleneaminopropylene (CH2CH 2

NHCH

2

CH

2

CH

2 and ethylenethioethylene (CH2CH 2

SCH

2

CH

2 The nitrogen atom interrupted in the alkylene group is optionally substituted by a lower alkyl group. The lower alkyl group is a C1- 4 alkyl group, of which a methyl group is preferred.

Examples of the "alkylene group which is optionally interrupted by one oxygen, itrogen or sulfur atom" include ethylene, trimethylene, tetramethylene, Doc: 97162p2.doc

P

7 9 5 6 9 IFP-9716(PyCT)/tsa -i/nglish translation/07. 10.98 pentamethylene, ethyleneoxyethyl, ethylenethioethyl, ethyleneaminoethyl, ethyleneaminopropyl, ethylene(methylamino)ethyl, ethylene(ethylamino)ethyl and ethylene(methylamino)propyl groups; of which a trimethylene, tetramethylene, pentamethylene, ethyleneoxyethyl, ethylenethioethyl, ethyleneaminoethyl, ethylene(methyl)aminoethyl or ethyleneaminopropyl group is preferred, a tetramethylene group being more preferred.

Preferred examples ofR 1 include a hydrogen atom and a C 1 4 alkyl group; of which a hydrogen atom or a methyl or ethyl group is more preferred, a hydrogen atom or a methyl group being most preferred.

Preferred examples ofR 2 include a hydrogen atom and a C 1 4 alkyl group; of which a hydrogen atom or a methyl or ethyl group is more preferred, a hydrogen atom or a methyl group being most preferred.

Preferred examples ofR 3 include a hydrogen atom, a CI-4 alkyl group, a C 1 -4 alkyl group having 1 to 3 substituents (examples of said substituent include a hydroxyl group, halogen atoms, a carbamoyl group, a carbamoyl group substituted with 1 or 2

C

1 -4 alkyl groups, a carbamoyloxy group, a carbamoyloxy group substituted with 1 or 2

C

14 alkyl groups, a C 1 4 alkoxy group, an amino group and an amino group substituted with 1 or 2 C-4 alkyl groups), a C 3 -6 cycloalkyl group and a group of formula

C(=NHR

4 )(wherein R 4 represents a hydrogen atom, a C 1 -4 alkyl group or an amino group); of which a hydrogen atom or a methyl, ethyl, 2 -hydroxyethyl, 2-fluoroethyl, 2 ,2,2-trifluoroethyl, carbamoylmethyl, 2 -carbamoylethyl, N-methylcarbamoylmethyl, N,N-dimethylcarbamoylmethyl, 2 -methoxyethyl, 2 -carbamoyloxyethyl, 2-aminoethyl, 2 -(methylamino)ethyl, 2 -(dimethylamino)ethyl, cyclopropyl, formimidoyl, acetimidoyl or amidino group is preferred, a hydrogen atom or a methyl, 2-hydroxyethyl, carbamoylmethyl, 2-aminoethyl, 2 -(methylamino)ethyl, cyclopropyl, formimidoyl, acetimidoyl or amidino group being more preferred, and a hydrogen atom or a methyl, formimidoyl, acetimidoyl or amidino group being most preferred.

Preferred examples of the group of R 2 and R 3 taken together include a C 2 6 Salkylene group which is optionally interrupted by one oxygen, nitrogen or sulfur atom S(said nitrogen atom is optionally substituted by a C 1 -4 alkyl group); of which a Doc: 97162p2.doc

P

79 5 69/FP-9716(PCTjytsa-ig/Engijsh translation/07.10.98 trimethylene, tetramethylene, pentamethylene, ethyleneoxyethylene, ethylenethioethylene, ethyleneaminoethylene, ethylene(methylamino)ethylene or ethyleneaminopropylene group is preferred, a tetramethylene group being most preferred.

The compound can be converted into its "pharmacologically acceptable salts or derivatives" if necessary.

Examples of the "pharmacologically acceptable salts" include salts of a mineral acid such as hydrochloride, hydrobromide, hydroiodide, phosphate, sulfate and nitrate; sulfonates such as methanesulfonate, ethanesulfonate, benzenesulfonate and ptoluenesulfonate; organic acid salts such as oxalate, tartrate, citrate, maleate, succinate, acetate, benzoate, mandelate, ascorbate, lactate, gluconate and malate; amino acid salts such as glycine salt, lysine salt, arginine salt, ornithine salt, glutamate and aspartate; inorganic salts such as lithium salt, sodium salt, potassium salt, calcium salt and magnesium salt; and salts with an organic base such as ammonium salt, triethylamine salt, diisopropylamine salt and cyclohexylamine salt; of which a hydrochloride, hydrobromide, phosphate, sulfate, methanesulfonate, p-toluenesulfonate, oxalate, tartrate, citrate, acetate, lactate, glutamate, aspartate, sodium salt, potassium salt, ammonium salt or triethylamine salt is preferred, a hydrochloride, sulfate, methanesulfonate, citrate, acetate or lactate being more preferred, and a hydrochloride or sulfate being most preferred.

The compound of the present invention happens to absorb water and form a product with absorbed water or a hydrate when it is left alone in the air, prepared by the lyophilization of its aqueous solution, or recrystallized. Such salts are also included in the present invention.

The "pharmacologically acceptable derivative" is a derivative in which some of the carboxyl, hydroxyl and amino groups or the like of compound are protected by a protecting group (a so-called prodrug-forming group) which may be cleaved in vivo by a chemical or biological method such as hydrolysis to afford the original compound

(I)

or salt thereof. Whether a derivative is such a derivative or not can be determined by orally or intravenously administering the derivative to an animal such as rat or mouse 97162p2.doc

P

79

S

6 9 /F?9716(PCrytsa-ig/Engijsh translation/07.1 0.98 and studying the body fluid of the animal. If the original compound or a pharmacologically acceptable salt thereof can be detected from the body fluid, the derivative is determined as a prodrug of compound Examples of such a protecting group for the carboxyl, hydroxyl, amino groups or the like include acyloxyalkyl, alkoxycarbonyloxyalkyl, phthalidyl, 2 -oxo-1,3-dioxolen-4-yl)alkyl which may have an alkyl or aryl group at the 5-position, acyl, alkoxycarbonyl and aminoacyl groups.

Examples of the acyloxyalkyl group include pivaloyloxymethyl, isobutyryloxymethyl, 1-(isobutyryloxy)ethyl, acetoxymethyl, 1-(acetoxy)ethyl, 1methylcyclohexylcarbonyloxymethyl, 1-methylcyclopentylcarbonyloxymethyl, 2ethylbutyryloxymethyl and hexanoyloxymethyl groups; of which a pivaloyloxymethyl, acetoxymethyl or 1-methylcyclohexylcarbonyloxymethyl group is preferred.

Examples of the alkoxycarbonyloxyalkyl group include tbutoxycarbonyloxymethyl, I1-(methoxycarbonyloxy)ethyl, 1-(ethoxycarbonyloxy)ethyl, 1-(isopropoxycarbonyloxy)ethyl, 1-(t-butoxycarbonyloxy)ethyl, 1- (cyclohexylcarbonyloxy)ethyl and 1-(cyclopentylcarbonyloxy)ethyl groups; of which a 1-(isopropoxycarbonyloxy)ethyl or 1-(cyclopentylcarbonyloxy)ethyl group is preferred.

Examples of the 1-( 2 -oxo-1,3-dioxolen-4-yl)alkyl group which may have an alkyl or an aryl group at the 5-position include 2 -oxo-1, 3 -dioxolen-4-ylmethyl, 1-(2oxo-1,3-dioxolen-4-yl)ethyl, 5-methyl-2-oxo-1,3-dioxolen-4-ylmethyl, 1-(5-methyl-2oxo-1,3-dioxolen-4-yl)ethyl, 5-ethyl-2-oxo-1, 3 -dioxolen-4-ylmethyl, 5-propyl-2-oxo- 1,3-dioxolen-4-ylmethyl and 5-phenyl-2-oxo-1,3-dioxolen-4-ylmethyl groups; of which a 5-methyl-2-oxo-1,3-dioxolen-4-ylmethy group is preferred.

Among the above-exemplified protecting groups, a 5-methyl-2-oxo-1,3dioxolen-4-ylmethyl, acetoxymethyl, pivaloyloxymethyl, 1methylcyclohexylcarbonyloxymethyl, 1-(isopropoxycarbonyloxy)ethyl or 1- (cyclohexyloxycarbonyloxy)ethyl group is more preferred and they are preferably used as a protecting group of the carboxyl group to make an ester derivative of compound The compounds of the present invention include individual isomers and a mixture of the isomers. The preferred example of the isomer is a compound which has an R configuration at the 1-position of the carbapenem skeleton, a (5S,6S) configuration Doc: 97162p2.doc

P

7 9 5 6 9 /FP-9716(PCI)/tsa-ig/English translation/07.10.98 at the 5- and 6-positions similarly to thienamycin, and an R configuration as a hydroxylcontaining -carbon at the substituent of the 6 -position.

The (2S,4S) configuration is suited for the 2- and 4-positions of the 2- (substituted pyrrolidine)-4-ylthio group of the substituent at the 2-position of the carbapenem skeleton.

There is no particular limitation on the configuration at the 3 -position of the 3- (substituted aminomethyl)pyrrolidin-l-yl group.

Preferred examples of the compound of formula include the compounds wherein:

R

1 represents a hydrogen atom or a C 1 4 alkyl group;

R

2 represents a hydrogen atom or a C 1 4 alkyl group; and

R

3 represents a hydrogen atom, a CI- 4 alkyl group, a CI-4 alkyl group having 1 to 3 substituents (each of said substituent represents a hydroxyl group, a halogen atom, a carbamoyl group, a carbamoyl group substituted by 1 or 2 Ci- 4 alkyl groups, a carbamoyloxy group, a carbamoyloxy group substituted by 1 or 2 C 1 -4 alkyl groups, a

C

1 4 alkoxy group, an amino group or an amino group substituted by 1 or 2 C1- 4 alkyl groups), a C3- 6 cycloalkyl group or a group of formula -C(=NH)R 4 (wherein

R

4 represents a hydrogen atom, a C 1 4 alkyl group or an amino group), or

R

2 and R 3 taken together represent a C 2 6 alkylene group which is optionally interrupted by one oxygen, nitrogen or sulfur atom (said nitrogen atom may be substituted by the C 1 4 alkyl group).

More preferred examples include compounds wherein:

R

1 represents a hydrogen atom or a methyl or ethyl group; R represents a hydrogen atom or a methyl or ethyl group; and

R

3 represents a hydrogen atom or a methyl, ethyl, 2-hydroxyethyl, 2-fluoroethyl, 2 2 ,2-trifluoroethyl, carbamoylmethyl, 2 -carbamoylethyl, N-methylcarbamoylmethyl, N,N-dimethylcarbamoylmethyl, 2 -methoxyethyl, 2-carbamoyloxyethyl, 2-aminoethyl,

P

7 9 56 9 /FP.9716(PCT)/tsa-ig/English translation/07.1 0.98 2 -(methylamino)ethyl, 2-(dimethylamino)ethyl, cyclopropyl, formimidoyl, acetimidoyl or amidino group, or

R

2 and R 3 taken together represent a trimethylene, tetramethylene, pentamethylene, ethyleneoxyethylene, ethylenethioethylene, ethyleneaminoethylene, ethylene(methylamino)ethylene or ethyleneaminopropylene group.

Still more preferred examples include compounds wherein: R represents a hydrogen atom or a methyl group;

R

2 represents a hydrogen atom or a methyl group; and

R

3 represents a methyl, 2-hydroxyethyl, carbamoylmethyl, 2-aminoethyl, 2- (methylamino)ethyl, cyclopropyl, formimidoyl, acetimidoyl or amidino group, or

R

2 and R3 taken together represent a tetramethylene group.

Most preferred examples include compounds wherein: R' represents a hydrogen atom or a methyl group;

R

2 represents a hydrogen atom or a methyl group; and

R

3 represents a hydrogen atom or a methyl, formimidoyl, acetimidoyl or amidino group.

In the above-exemplified preferred, more preferred, still more preferred and most preferred compounds the stereoisomer represented by the following formula: OH

H

3 CON NR2 H H C

ON,<

S R3

CH

3 S /N R1 0

COOH

or the following formula: Doc: 97162p2.doc

P

7 9 5 6 9/FP-9716(PCT)/tsa-ig/English translation/07.10.98 0 -N-ICOOH NR1 are more preferred compounds.

The preferred compounds of the formula can be exemplified in Table 1.

[Table 1] OH CH 3

CONNR

COH

3 S :TR 0

COOH

Exemplified R1 R 2

R'

Compound No.

1 H H Me 2 H H Et 3 H H CH 2

CH

2

OH

4 H H CH 2

CH

2

F

H H CH1 2

CF

3 6 H H CH 2

CONH

2 7 H H CH 2

CH

2

CONH-

2 8 H H CH 2 CONHMe 9 H H CH 2 CONMe 2 H H CH 2

CH

2 OMe, 97162p2.doc 971622.docP 7 9 5 6 9 /FP-9716(PCTyta-ig/English translation/07. 10.98 11 H H CH 2

CH

2 OCONHj 2 12 H H CH 2

CH

2

NH

2 13 H H CH 2

CH

2 NHMe 14 H H CH 2

CH

2 NMe 2 H H cOr 16 H H C(=NH)H 17 H H C(=NH)Me 18 H H

C(=NH)NH

2 19 H Me Me H Me Et 21 H Me CH 2

CH

2

OH

22 H Me CH 2

CH

2

F

23 H Me CH 2

CF

3 24 H Me

CH

2

CONH

2 H Me -CH 2

CH

2

CONH

2 26 H Me CH 2 CONIIMe 27 H Me

CH

2 CONMe 2 28 H Me CH 2

CH

2 OMe 29 H Me

CH

2

CH

2

OCONH

2 H Me

CH

2

CH

2

NH

2 31 H Me CH 2

CH

2 NHMe 32 H Me

CH

2

CH

2 NMe 2 33 H Me O~r 34 H Me C(=NH)H H Me C(=NH)Me Doc: 97162p2.doc Doc: 7162p.docP 7 9 5 6 9IFP-9716(PCT)/tsa-grEnlgish t-anslationO7. 10.98 36 H Me C(=NIJ)NH 2 37 H Et Et 38 H Et CH 2

CH

2

OH

39 H Et CH 2

CH

2

F

H Et CH 2

CF

3 41 H Et CH 2

CONHF

2 42 H Et CH 2

CH

2

CONH

2 43 H Et CH 2 CONII~e 44 H Et CH 2 CONMe 2 H Et CH 2

CH

2 OMe 46 H Et CH 2

CH

2

OCONII

2 47 H Et CH 2

CH

2

NH

2 48 H Et CH2CH 2 NHMe 49 H Et CH 2

CH

2 NMe 2 H Et O~r 51 H Et C(=NH)H 52 H Et C(=NH)Me 53 H Et C(=NH)NH 2 54 H -CH 2

CH

2

CH

2

CH

2 H -CH2CH 2

CH

2

CH

2

CH

2 56 H -CH 2

CH

2

OCH

2

CH

2 57 H -CH 2

CH

2

SCH

2

CH

2 58 H -CH 2

CH

2

NHCH

2

CH

2 59 H -CH 2

CH

2

NHCH

2

CH

2

CH

2 H -CH2CH 2 MeCH 2

CH

2 Doc: 97162p2.doc Doc:9712p2docP 79 56 9 /FP-9716(PCT)/tsa-ig/Engijsh translation/O7.1O.98 61 62 63 64 66 67 68 69 71 72 73 74 76 77 78 79 81 82 83 84 Me H Me H Me H Me H Me H Me H Me H Me H Me H Me H Me H Me H Me H Me H Me H Me H Me H Me H Me H Me Me Me Me Me Me Me Me Me Me Me Me

H

Me Et

CH

2

CH

2

OH

CH

2

CH

2

F

CH

2

CF

3

CH

2

CONI{

2

CH

2

CH

2

CONH

2

CH

2 CONHIMe

CH

2 CONMe 2

CH

2

CH

2 OMe CH2CH 2

OCONH

2

CH

2

CH

2

NH

2

CH

2

CH

2 NHMe

CH

2

CH

2 NMe 2 cOr

C(=NH)H

C(=NH)Me

C(=NH)NH

2 Me Et

CH

2

CH

2

OH

CH

2

CH

2

F

CH

2

CF

3

CH

2

CONII

2 Doc: 97162p2.doc Doc: 7162p.docP 7 9 569/FP-9716(PC'Iytsa.ig/Engijsh translationO7. 10.98 86 87 88 88 89 91 92 93 94 96 97 98 99 100 101 102 103 104 105 106 107 108 109 Me Me Me Me Me Me Me Me Me Me Me Me Me Me Me Me Me Me Me Me Me Me Me Me Me Et Me Et Me Et Me Et Me Et Me Et Me Et Me Et Me Et Me Et Me Et Me Et Me Et

CH

2

CH

2

CONFI

2

CH

2 CONHiMe

CH

2 CONMe 2

CH

2

CH

2 OMe

CH

2

CH

2

OCONII

2

CH

2

CJJ

2

NH

2

CH

2

CL{

2 NH~ve

CH

2

CH

2 NMe 2 O~r

C(=NH)H

C(=NH)Me

C(=NHW)H

2 Et

CH

2

CH

2

OH

CH

2

CH

2

F

CH

2

CF

3

CH

2

CONH

2

CH

2

CH

2

CONH

2

CH

2 CONHMe,

CH

2 CONMe 2

CH

2

CH

2 OMe

CH

2

CH

2

OCONH

2

CH

2

CH

2

NI{

2

CH

2

CH

2 NMe

CH

2

CH

2 NMe 2 Doe: 97162p2.doc Doc: 7162p.docP 7 9 5 6 9IFP-9716(PCT)/tsa-ipg/English translationio7. 10.98 110

III

112 113 114 115 116 117 118 119 120 121 122 123 124 125 126 127 128 129 130 131 132 133 134 Me Me Me Me Me Me Me Me Me Me Me Et Et Et Et Et Et Et Et Et Et Et Et Et Et Et O~r Et C(=NH)H Et C(=NH)Me Et

C(=NII)NH

2

-CH

2

CH

2

CH

2

CH

2

-CH

2

CH

2

CH

2

CH

2

CH

2

-CH

2

CH

2

OCH

2

CH

2

-CH

2

CH

2

SCH

2

CH

2

-CH

2

CH

2

NHCH

2

CH

2 -CH2CH 2

NHCH

2

CH

2

CH

2 -CH2CH 2 NMeCH 2

CH

2 H H H Me H Et H CH 2

CH

2

OH

H CH 2

CH

2

F

H

CH

2

CF

3 H

CH

2

CONH

2 H

CH

2

CH

2

CONH

2 H CH 2 CONHMe H

CH

2 CONMe 2 H

CH

2

CH

2 OMe H

CH

2

CH

2

OCONH

2 H

CH

2

CH

2

NH

2 H CH 2

CH

2 N~Hve, Doc: 97162p2.doe Doc: 7162p.docP 7 9 5 6 9 /FP-9716(PCT)/tsa-ig/English translationO7. 10.98 135 Et H CH 2

CH

2 NMe 2 136 Et H O~r 137 Et H C(=NH)H 138 Et H C(=NH)Me 139 Et H C(=NH)NH 2 140 Et Me Me 141 Et Me Et 142 Et Me CH 2

CH

2

OH

143 Et Me CH 2

CH

2

F

144 Et Me CH 2

CF

3 145 Et Me CH 2

CONH

2 146 Et Me CH 2

CH

2

CONH

2 147 Et Me CH 2 CONHMe 148 Et Me CH 2 CONMe 2 149 Et Me CH 2

CH

2 OMe 150 Et Me CH 2

CH

2

OCONH

2 151 Et Me CH 2

CH

2

NH

2 152 Et Me CH 2

CH

2 NHMe 153 Et Me CH 2

CH

2 N'Me 2 154 Et Me cOr 155 Et Me C(=NH)H 156 Et Me C(=NH)Me 157 Et Me C(=NH)NH 2 158 Et Et Et V~9 Et Et CH 2

CH

2

OH

Doc: 97162p2.doc Dac: 7162p.docP 7 9 5 6 9 /FP-9716(PCT)tsaigEnglish trnnslationO7. 10.98 160 Et Et CH 2

CH

2

F

161 Et Et CH 2

CF

3 162 Et Et CH 2

CONII

2 163 Et Et CH 2

CH

2

CONH

2 164 Et Et CH 2 CONHNe 165 Et Et CH 2 CONMe 2 166 Et Et CH 2

CH

2 OMe 167 Et Et CH 2

CH

2

OCONH

2 168 Et Et CH 2

CH

2

NH

169 Et Et CH2CH 2 NEJMe 170 Et Et CH 2

CH

2 NMe 2 171 Et Et O~r 172 Et Et C(=NH)H 173 Et Et C(=NH)Me 174 Et Et C(=NH)NH 2 175 Et -CH 2

CH

2

CH

2

CH

2 176 Et -CH2CH 2

CH

2

CH

2

CH

2 177 Et -CH 2

CH

2

OCH

2

CH

2 178 Et -CH 2

CH

2

SCH

2

CH

2 179 Et -CH 2

CH

2

NHCH

2

CH

2 180 Et -CH 2

CH

2

NHCH

2

CH

2

CH

2 181 Et -CH 2

CH

2 NMeCH 2

CH

2 Doc: 97162p2.doe Doc: 7162p.docP 7 9 5 6 9 /FP-9716(PCI)/tsa-ig/English translationO7. 10.98 In the above Table-i, Me, Et and cPr represent a methyl, ethyl and cyclopropyl group, respectively.

Among the compounds exemplified in the above Table-i, compounds of Exemplified Compound Nos: 1, 3, 6, 12, 13, 16, 17, 18, 34, 35, 36, 61, 62, 77, 78, 79, 94, 95 and 96 are more preferred.

Of which, the compounds specified by the following chemical names are most preferred: Stereoisomers of Exemplified Compound No. 1: (1R, 5 S, iR)-l1-hydroxyethyl]-l1-methyl-2[(2S,4S)2[(3R)-3methylaminomethylpyrrolidin I -ylcarbonyl]pyrolidin4ylthioy 1 -carbapen-2-em-3 carboxylic acid (Compound of Example 3) (1R, 5 S, 1R)-l1-hydroxyethyl]-l1-methyl-2-[(2S,4S)2[(3 S)-3 methylaminomethylpyffolidin-1 -ylcarbonyl]pyrolidin4ylthioy 1 -carbapen-2-em-3 carboxylic acid (Compound of Example Stereoisomers of Exemplified Compound No. 3: (1 R, 5S, 1R)-l1-hydroxyethyly2-[(2 hydroxyethylaminomethyl)pyrrolidin-1 -ylcarbonyl]- 1-methyl-i -carbapen-2-em-3carboxylic acid (1R, 5 IR)-l1-hydroxyethyl]-2-[(2S,4S)-2..{(3 S)-3 hydroxyethylaminomethyl)pyrrolidin I -ylcarbonyl]- 1-methyl-i -carbapen-2-em-3carboxylic acid (Compound of Example 13) Stereoisomers of Exemplified Compound No. 6: (I 5SS22S )2[3R--cramymtyamnmt1~pfoiiylcarbonyl]pyffolidin4ylthio-6-[( iR)- 1-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 carboxylic acid (Compound of Example 14)

P

7 9 5 6 9 IFP-9716(PCT)ts.ig/English tasjaionO7. 10.98 (I R,

S)-

3 -(car-barnioylmicthiylamiinoniethyl)pyrrolidin 1ylcarboniyllpyrr-olidin-4.yltio-6-[( I I -hydroxyethyl]- 1 -methyl- I -carbapen-2-em- 3-carboxylic acid Stercoisomer-s of Exemplified Compound No. 12:- (1 R, 5

S,

6 2 S,4S)-2-[(3R)-3-(2.ain iinoethylamijinmthiyl)pyn-olidinyl carbonyl]pyrrol id in-4.ylthio]-6-[( I I -hydroxyethyl]- I -m ethyl- I -carbapen-2-em- 3-carboxylic acid (1 R, 5 S, S, S)-3 2 -am inoethylaminmom ethyl)pyrrol idin- 1ylcarbonyl] pyrrOlI Id In-4-yl thio.-6{(I 1 -hydroxyeth yl]-I -m ethyl- I -carbapen-2-em- 3-carboxylic acid Stereoisomers of Exemplified Compound No, 13: (I R, 5 2 -N-methylam i no)ethylam m omn ethyl)pyrrolidin-1I-ylcarbonyl] pyrrolidin-4-ylthio]-6-[( IR)-lI-hydroxyethyl]- 1-methyl-I carbapen-2-em-3-carboxylic acid (I R, 5 S, S,4 [3 S)-3 -(2-N-rnethlylain ino)ethyl am i nomnethyl)pyrrolidin-1I-ylcarbonyl] pyrroli1din-4-ylth io]-6 IR)- I -hydroxyethyl]-I 1-mnethyl- I :carbapen-2-em-3-carboxylic acid V Stercoisoniers of Exemplified Compound No. 16.- (I R, 5 ,6)2[2S4S--(R- fr i dy minmehlyrlidn I 0 0 0 y IcarbonylIipyrro II d i n-4-ylth io-6[+ 1 -h ydroxyethyl ]-1I-miethyl -I -carbapen-2-ern- 3-carboxylic acid (Compound of Example 23) 0 (1 R, 5

S)-

3 -formimidoylam inomnethylpyrroli din- 1- 0 yl carbonyl ]pyrrol di n4ylth 1o] R)-1I -hydroxyethyl]- 1 -m ethyl -I -carbapen-2-em 3-carboxylic acid o Stereoisomers of Exemplified Compound No. 17: (1R,5 S, 6

S)-

2 -[(2S,4S)-2-(3R)-3-acetimidoylammnomethylpyrroIjdi I yI carbonyl ]pyrrol 1di n4-ylth ioy&6[( I I -hydroxyethyl]- I -m ethyl -I -carbapen-2-em 3-carboxylic acid (Compound of Example 8) (1 R, 5 S, S)-3 -acetimidoylaminomethylpyn-olidin 1ylcarbonyl]pyrrolidin-4-yltffioy6-[( IR)- 1 -hydroxyethyl]- 1-methyl- I -carbapen-2-em-3carboxylic acid (Compound of Example Stereoisomers of Exemplified Compound No. 18: (1 R, 5S, 6

S)-

2 2 S,4S)-2-[(3R).3.giuanidinomethylpyrrolid in 1 yl carbonyl]pyrrolidin4.ylthio]6[( 1 1 -hydroxyethyl]-I 1-m ethyl- I -carbapen-2-em-3 carboxylic acid (Compound of Example 7) (I R, 5 S, 6 S, 4 S)-3 -guanidinomethylpyn-oli din- 1ylcarbonyl]pyrrolidin-4.ylthio-6-[( 1R)-l1-hydroxyethyl]- 1-methyl-I -carbapen-2-em-3 carboxylic acid (Compound of Example 11) Stereoisomers of Exemplified Compound No. 34: formnimidoylaminomethylpyrrolidin- 1 -ylcarbonyl]pyrrolidin4ylthio-6-[( 1R)- 1 hydroxyethyl]- 1 -methyl- I -carbapen-2-em-3-.carboxylic acid (Compound of Example 21) (1 R, 5 S, S)-3 -N-methyl-Nformimi doylaminomethylpyr-olidin- 1 -ylcarbonyl ipyrrolidin-4-ylthio]-6[( 1 1 hydroxyethyl]- 1 -mnethyl- I -carbapen-2-em-3 -carboxylic acid Stereoisomers of Exemplified Compound No. (1 R, 5S, 6

S)-

2 (2S,4S)2[(3R).3N..methyl.N acetimidoylaminomethylpyrrolidin-l-ylcarbonyl]pyrrolidin4ylthio-6-[( 1R)- 1hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 -carboxylic acid (Compound of Example (iR 5 S)-3 -N-methyl-Nacetimidoylaminomethylpyrrolidin-l-ylcarbonyl]pyrrolidin-4ylthio-6-[( 1R)- 1hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 -carboxylic acid 97162pldoc 97162p2.docP 7 9 569/FP-9716(PCI)/tsa-igEnglish tz-aslation/07. 10.98 Stereoisomers of Exemplified Compound No. 36: 1 -methylguanidinomethyl)pyrroidin-

I-

ylcarbonyl]pyrrolidin-4-ylthio]-6[( 1R)- 1 -hydroxyethyl]- 1 -methyl- I -carbapen-2-em-3 carboxylic acid (Compound of Example 19) (1 R,5 S, S)-3 -methylguanidinomethyl)pyrrol idin- 1ylcarbonyl]pyrrolidin-4-ylthio]-6[( IR)-l1-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 carboxylic acid Stereoisomers of Exemplified Compound No. 61: (1 R, 5 S, 6 S, 4 -am inomethylpyrroli din- 1 -ylcarbonyl] 1methylpyrrolidin-4-ylthio-6-[( 1R)-lI-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 carboxylic acid (Compound of Example 1) (1 R, S)-3 -aminomethylpyrrolidin-1 -ylcarbonyl]- 1methylpyrrolidin-4-ylthio]-6[( iR)-lI-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 carboxylic acid (Compound of Example 2) Stereoisomers of Exemplified Compound No. 62: (1 R, 1R)-l1-hydroxyethyl]-l1-methyl-2-[(2S,4S)2-[(3R)-3methylaminomethylpyrr.olidin-1 -ylcarbonyl]-l1-methylpyrrolidin-4-ylthioy 1 -carbapen- 2 -em-3-carboxylic acid (Compound of Example 4) (i iR)-lI-bydroxyethyl]- 1-methyl-2-[(2S,4S)-2-[(3 S)-3methylaminomethylpyrrolidin I -ylcarbonyly I -methylpyrrolidin-4-ylthioy 1 -carbapen- 2 -em-3-carboxylic acid (Compound of Example 6) Stereoisomers of Exemplified Compound No. 77: (1 5SS22S )2[3R--omiiolmioeh1pfoiiylcarbonyl]-l1-methylpyrrolidin-4ylthio]6[( 1R)-l1-hydroxyethyl]- 1-methyl-i carbapen-2-em-3-carboxylic acid (Compound of Example 26) Doe: 97162p2.doc Doc: 7162p.docP 7 9 5 6 9fFP-9716(PCT)/tsa-ig/English translation/O7. 10.98 (iR,

S)-

3 -form imidoylamninomethylpyrroli din- 1ylcarbonyl]- 1 -methylpyrroiidin-4-ylthio-6-[( iR)- 1 -hydroxyethyl]- 1 -methyl- I carbapen-2-em-3-carboxylic acid Stereoisomers of Exemplified Compound No. 78: (1 R, 5 S, 6 S,4 3

R)-

3 -acetimidoylaminom ethylpyrrol idin- 1ylcarbonyl]- 1 -methylpyrrolidin-4-ylthio-6-[( iR)- 1 -hydroxyethyl]- 1 -methyl- I carbapen-2-em-3 -carboxylic acid (Compound of Example (1 R, 5 S, 6S)-2-[(2S,4 S)-3 -acetimidoylaminomethylpyn-oiidin 1ylcarbonylj- 1 -methylpyrrol idin-4-ylthio]-6-[( 1R)- 1 -hydroxyethyl]- 1 -methyl- I carbapen-2-em-3-carboxylic acid Stereoisomers of Exemplified Compound No. 79: (1 R, SS, 6

S)-

2 [(2S,4S)-2..{(3R)-3..gianidinomethylpyrrolidi 1 -ylcarbonyl]- 1m ethylpyrrolidin-4-ylthio 1 1 -hydroxyethyl]- 1 -m ethyl- I -carbapen-2-em-3 carboxylic acid (Compound of Example 24) (I R, 5 S, 6 S,4 S)-3 -guanidinom ethylpyrrol idin- 1 -ylcarbonyl]- 1 methylpyrrolidin-4-ylthio]6[(1R)-1-hydroxyethyl]- 1-methyl-I -carbapen-2-em-3carboxylic acid Stereoisomers of Exemplified Compound No. 94: (1R, 5

S,

4 S)-2-[(3R)-3-N-methyl.N.

formimidoylaminomethylpyr-olidin. 1-ylcarbonyl]-l1-methylpyrrolidin-4ylthio.6- R)-l1-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3-carboxylic acid (Compound of Example 29) (1R, 5 S, S)-3 -N-methyl-Nformnimidoylaminomethylpyrrolidin-1 -ylcarbonyl]-l1-methylpyrrolidin-4..ylthioy-6 [(iR)-lI-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 -carboxylic acid P79569/FP-9716(PCTytsa-ig/English translation/07. 10.98 Stereoisomers of Exemplified Compound No.

S,

6

S)-

2 -E(2S,4S).2[(3R).3.Nmethyl.N acetimidoylaminomethylpyrroli din- 1 -ylcarbonyl]-I 1i methylpyrrolidin-4-ylthio]-6-[( 1 R)- 1 -hydroxyethyl]-I 1-m ethyl -I -carb apen-2-em-3 -carboxyli c acid (Compound of Example 28) S)-3-N-methyl-Nacetimidoylaminomethylpyrroli din- I -ylcarbonyl]- I -methylpyrrolidin-4-ylthio-6-[( 1 R)- 1 -hydroxyethyl]-I 1-m ethyl- I -carbapen-2-em-3 -carboxylic acid Stereoisomers of Exemplified Compound No. 96: (1 R, 5S, S,4 1-methylguanidinomethyl)pyrrolidinylcarbonyl]-l1-methylpyrrolidin-4-ylthio-6-[( 1R)-l1-hydroxyethyl]- 1-methyl-i carbapen-2-em-3-carboxylic acid (Compound of Example 27) (1 R 5 S, S)-3 -methylguanidinomethyl)pyn-olidin i ylcarbonyl]-l1-methylpyrrolidin-4-ylthio-6-[( iR)-l1-hydroxyethyl]- i-methyl-i carbapen-2-em-3-carboxylic acid The i -methylcarbapenem derivatives of the present invention represented by the formula can be prepared by reacting a carbapenem compound of formula (11): OH

CH

3

CH

3

IRL

O

COOR

(wherein R L represents a leaving group and R 5 represents a carboxy protecting group) with a mercaptopyrrolidine derivative of formula (111): CONW p H S -CRN p N'R1 Doe: 97162p2.doe Doc: 7162p.docP 7 9 5 69IFP-9716(PCT1,ta.jgEnglish traslationO7. 10.98 (wherein R'p represents an amino protecting group or has the same meaning as described in R 1

R

2 p represents an amino protecting group or has the same meaning as described in R 2 except that the amino, hydroxyl or imino group contained in R 2 may be protected, R 3 p represents an amino protecting group or has the same meaning as described in R 3 except that the amino, hydroxyl or imino group contained in R 3 may be protected) and then removal of the protecting group if necessary. Furthermore, it can be converted into its pharmacologically acceptable salts or derivatives if necessary.

The compound of the present invention can be prepared by processes (Method A or Method B) illustrated below.

P79569/FP-9716(PCTtsa-ig/English translation/07. 10.98 (Method A) O H

OH

3 0 0

COORS

(IV)

Step Al

~R

Step A2 CONO

I~

HS NN OH C Step A3

OH

3 0 (Method B)

CH

3

ONO

O~a N IR

COOH

(I)

cw.I Step Bi

CH

3 S(O)R7 HS-: "R3p OH CH 3 CON(NRp

OH

3 S

NR

3 p Step B2 0

(VI)

COOH

(I)

97162p2.doe 971622.docP 7 95691FP-97l6(PCTytsa.jg/English trnmslaionO7. 10.98 In the above formulae, R2, R3, Rip, R 2 p and R 3 p have the same meanings as described above, and R 5 represents a carboxy protecting group.

The protecting group of the hydroxyl, amino or imino group contained in R'p, R2p or R3p is a protecting group ordinarily used in the field of organic synthetic chemistry (Greene Wuts, Protective Groups in Organic Synthesis, 2nd Edition, 1991, John Wiley Sons, Inc.). Preferred examples include a benzyl group which may have a substituent such as benzyl and 4-nitrobenzyl; a benzyloxycarbonyl group which may have a substituent such as benzyloxycarbonyl and 4 -nitrobenzyloxycarbonyl; or an allyloxycarbonyl group which may be substituted at the 2-position such as allyloxycarbonyl, 2 -chloroallyloxycarbonyl and 2 -methylallyloxycarbonyl; of which a 4-nitrobenzyl or 4 -nitrobenzyloxycarbonyl group is more preferred.

The "carboxy protecting group" as R5 is a protective group ordinarily used in the field of synthetic organic synthesis (Greene Wuts, Protective Groups in Organic Synthesis, 2nd Edition, 1991, John Wiley Sons, Inc.). Examples include a C 14 alkyl group such as methyl, ethyl or t-butyl; a benzyl group which may have a substituent such as benzyl, 4-methoxybenzyl, 4-nitrobenzyl or 2-nitrobenzyl; a benzhydryl group; an allyl group which may have a substituent at the 2-position such as allyl, 2-chloroallyl or 2-methylallyl; a halogenoethyl group such as 2 2 ,2-trichloroethyl, 2,2-dibromoethyl or 2,2,2-tribromoethyl; or 2-trimethylsilylethyl group; of which a 4-nitrobenzyl or benzyl group is more preferred.

R6 represents a C 1 4 alkanesulfonyl group such as methanesulfonyl, trifluoromethanesulfonyl, ethanesulfonyl, propanesulfonyl, isopropanesulfonyl or butanesulfonyl; a C 6 -o 10 arylsulfonyl group such as phenylsulfonyl, tolylsulfonyl or naphthylsulfonyl; a di-(C. alkyl)phosphoryl group such as dimethylphosphoryl, diethylphosphoryl, dipropylphosphoryl, diisopropylphosphoryl, dibutylphosphoryl, dipentylphosphoryl or dihexylphosphoryl; or a di(Cs 6 -o aryl)phosphoryl group such as diphenylphosphoryl or ditolylphosphoryl; of which a diphenylphosphoryl group is preferred.

R7 represents a C- 4 alkyl group such as methyl, ethyl, propyl or isopropyl; a halogeno-(C- 1 4 alkyl) group such as fluoromethyl, chloromethyl, fluoroethyl, chloroethyl, fluoropropyl, difluoromethyl, difluoroethyl, dichloroethyl, trifluoromethyl Doc: 97162p2.doc P79569/FP-9716(PCTy/tsa-igEngish translation/07. 10.98 or trifluoroethyl; a 2 -acetylaminoethyl group; a 2 -acetylaminovinyl group; a C6- 10 aryl group, such as phenyl or naphthyl, which may have substituents (said aryl group may have one to three substituents. They are the same as or different from each other and each substituent is described below. Examples include a halogen atom such as fluorine, chlorine and bromine; a C1-4 alkyl group such as methyl, ethyl, propyl and isopropyl; a

C

1 4 alkoxy group such as methoxy, ethoxy, propoxy and isopropoxy; a (CI-4 alkoxy)carbonyl group such as methoxycarbonyl, ethoxycarbonyl and t-butoxycarbonyl; a carbamoyl group, a mono- or di-(C1.

4 alkyl)carbamoyl group; a nitro group; a hydroxyl group; and a cyano group); or a heteroaryl group which has one or two nitrogen atoms, such as pyridyl or pyrimidinyl, and may have substituents (said heteroaryl group may have one to three substituents. They are the same or different from each other and each includes a halogen atom and a C 14 alkyl group which have been exemplified above as the substituent of the aryl group).

Incidentally, examples of the "leaving group" of RL include a group represented by formula -OR 6 or -S(O)R 7 In method A, a Compound is prepared by reacting the compound of formula (IV) with a sulfonylating or phosphorylating agent in the presence of a base to afford a compound of formula (Step Al); by reacting Compound with a compound of formula (III) in the presence of a base to give a compound of formula (VI) (Step A2); and finally, by removing any protecting groups from the compound of formula (VI) (Step A3). Each step will be described below.

(Step Al) In Step Al a compound of formula is prepared by reacting a compound of formula (IV) with a sulfonylating or phosphorylating agent in an inert solvent in the presence of a base.

Examples of the sulfonylating agent include C 1 4 alkanesulfonic anhydrides such as methanesulfonic anhydride, trifluoromethanesulfonic anhydride and ethanesulfonic Doc: 97162p2.doc

P

7 9569/FP-9716(PCTtsaig/English translation/07. 10.98 anhydride; and C-10o arylsulfonic anhydrides such as benzenesulfonic anhydride and ptoluenesulfonic anhydride; of which p-toluenesulfonic anhydride is preferred.

Examples of the phosphorylating agent include di(C 1 4 alkyl)phosphoryl halides such as dimethylphosphoryl chloride and diethylphosphoryl chloride; and di(C6-o1 aryl)phosphoryl halides such as diphenylphosphoryl chloride and diphenylphosphoryl bromide; of which diphenylphosphoryl chloride is preferred.

There is no particular limitation on the nature of the solvent to be employed, provided that it has no adverse effect on the reaction. Examples of suitable solvents include halogenated hydrocarbons such as methylene chloride, 1,2-dichloroethane and chloroform, nitriles such as acetonitrile, amides such as N,N-dimethylformamide and N,N-dimethylacetamide, esters such as ethyl acetate and methyl acetate, and ethers such as diethyl ether, tetrahydrofuran and dioxane; of which acetonitrile, N,Ndimethylformamide or tetrahydrofuran is preferred, acetonitrile being most preferred.

There is no particular limitation on the nature of the base to be employed, provided that it does not affect the other part of the compound, particularly the p-lactam ring. Preferred examples of the base include organic bases such as triethylamine, diisopropylethylamine, pyridine and 4 -dimethylaminopyridine; of which diisopropylethylamine is more preferred.

Although no particular limitation is imposed on the reaction temperature, reaction at a relatively low temperature is desirable in order to suppress side reactions.

The reaction is usually carried out at a temperature from -20 0 C to 40 0 C (preferably from 0 C to 20'C). The reaction time mainly depends on the reaction temperature or nature of reagents; however it ranges from 10 minutes to 5 hours (preferably from minutes to 1 hour).

After the completion of the reaction, a resulting compound of the present step is obtained from the reaction mixture by known means. For example, to the reaction mixture or to the residue obtained by distilling off the solvent from the reaction mixture, an organic solvent which is not miscible with water is added, followed by washing with water and distilling off the organic solvent. If necessary, the resulting compound can be further purified by known means, for example, by recrystallization, reprecipitation or chromatography. It is also possible to subject the resulting compound Doc: 97162p2.doc P79569/FP-971 6(PCT)/tsa-ig/English translation/07.10.98 to the subsequent reaction (step A2) without isolation from the reaction mixture, if desired.

(Step A2) In Step A2 a compound of formula (VI) is prepared by reacting a compound (V) with a mercaptopyrrolidine derivative of formula (III) in an inert solvent in the presence of a base.

There is no particular limitation on the nature of the solvent to be employed, provided that it has no adverse effect on the reaction. Examples of suitable solvents include halogenated hydrocarbons such as methylene chloride, 1, 2 -dichloroethane and chloroform; nitriles such as acetonitrile; amides such as N,N-dimethylformamide and N,N-dimethylacetamide; esters such as ethyl acetate and methyl acetate; and ethers such as diethyl ether, tetrahydrofuran and dioxane; of which acetonitrile,

N,N-

dimethylformamide or tetrahydrofuran is preferred, acetonitrile being more preferred.

Although no particular limitation is imposed on the nature of the base to be employed in the present step, preferred examples include organic amines such as triethylamine and diisopropylethylamine and inorganic bases such as potassium carbonate and sodium carbonate; of which diisopropylethylamine is more preferred.

Although no particular limitation is imposed on the reaction temperature, the reaction is usually carried out at a temperature from -20 0 C to 40 0 C (preferably from 0 C to 20 0 The reaction time ranges from 30 minutes to 108 hours (preferably from 1 hour to 18 hours).

After the completion of the reaction, the resulting compound (VI) of the present step is obtained from the reaction mixture by known means. For example, to the reaction mixture or to the residue obtained by distilling off the solvent from the reaction mixture, an organic solvent which is not miscible with water is added, followed by washing with water and distilling off the organic solvent. If necessary, the resulting compound can be further purified by known means, for example, by recrystallization, reprecipitation or chromatography. It is also possible to subject the resulting compound Doc: 97162p2.doc

P

7 9 5 6 9 /FP-9716(PCT)/tsa-ig/English translation/07.10.98 (VI) to the subsequent reaction (step A3) without isolation from the reaction mixture, if desired.

(Step A3) In Step A3, a compound (VI) is converted to a compound by removal of any protecting groups from the compound (VI).

Although the method for removal of a protecting group depends on the nature of it, the protecting group is usually removed by a method ordinarily employed in the field of synthetic organic chemistry (Greene Wuts, Protective Groups in Organic Synthesis, 2nd Edition, 1991, John Wiley Sons, Inc.).

When the protecting group R 5 is, for example, a benzyl group which may have a substituent, such as benzyl or 4-nitrobenzyl, or a benzhydryl group, and when the protecting group of the hydroxyl, amino or imino group contained in R'p, R 2 p or R 3 p is a benzyl group which may have a substituent, such as benzyl or 4-nitrobenzyl, or a benzyloxycarbonyl group which may have a substituent, such as benzyloxycarbonyl or 4 -nitrobenzyloxycarbonyl, the protecting group can be removed by reacting with a reducing agent such as the combination of hydrogen with a hydrogenation catalyst or an alkali metal sulfide.

Examples of the reducing agent include combinations of hydrogen with a hydrogenation catalyst such as palladium-carbon and alkali metal sulfides such as sodium sulfide and potassium sulfide; of which the combination of hydrogen with palladium-carbon is preferred.

There is no particular limitation on the nature of the solvent to be employed, provided that it has no adverse effect on the present reaction; however, alcohols such as methanol and ethanol, ethers such as tetrahydrofuran and dioxane and a mixture of said organic solvents and water are preferred.

The reaction temperature usually ranges from 0 C to 50 0 C (preferably from to 40 0 The reaction time depends on the natures of the starting compound and Doc: 97162p2.doc

P

7 9 56 9/FP-9716(PCT)/tsaig/English translation/07.10.98 the reducing agent; however it usually ranges from 5 minutes to 12 hours (preferably from 30 minutes to 4 hours).

After the completion of the reaction, a compound is obtained from the reaction mixture by known means. For example, the resulting compound can be obtained by filtering off an insoluble material from the reaction mixture and then distilling off the solvent.

When the protecting group R 5 is an allyl group which may be substituted at the 2-position such as allyl, 2-chloroallyl or 2-methylallyl and when the protective group of the hydroxyl group, amino group or imino group contained in R'p, R 2 p or R 3 p is an allyloxycarbonyl group which may be substituted at the 2-position such as allyloxycarbonyl, 2 -chloroallyloxycarbonyl or 2 -methylallyloxycarbonyl, the protecting groups can be removed by reacting with a deprotecting agent; for example, a palladiumtrialkyltin hydride such as bis(triphenylphosphine)palladium chloride-tributyltin hydride or tetrakis(triphenylphosphine)palladium-tributyltin hydride or a palladium-alkali metal salt of an organic carboxylic acid such as tetrakis(triphenylphosphine)palladiumpotassium 2-ethylhexanoate or -sodium 2-ethylhexanoate.

Preferred examples of the deprotecting agents include bis(triphenylphosphine)palladium chloride-tributyltin hydride and tetrakis(triphenylphosphine)palladium-potassium 2-ethylhexanoate.

There is no particular limitation on the nature of the solvent to be used, provided that it has no adverse effect on the present reaction. Examples include the halogenated hydrocarbons such as methylene chloride, chloroform and 1, 2 -dichloroethane, esters such as ethyl acetate, ethers such as tetrahydrofuran, dioxane and 1,2-dimethoxyethane, nitriles such as acetonitrile, alcohols such as methanol, ethanol and propanol and water, and a mixture thereof, of which methylene chloride, ethyl acetate and mixtures thereof are preferred.

Although no particular limitation is imposed on the reaction temperature, the reaction is usually carried out at a temperature from -20 0 C to 100 0 C (preferably from 0°C to 60°C). The reaction time usually ranges from 30 minutes to 48 hours (preferably from 30 minutes to 12 hours).

Doc: 97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English translation/07. 10.98 After completion of the reaction, a compound is obtained from the reaction mixture by known means. For example, the insoluble material precipitated by the reaction is filtered off from the reaction mixture, followed by distilling off the solvent, to afford a compound When the protecting group R 5 is a halogenoethyl group such as 2,2dibromoethyl or 2 2 ,2-trichloroethyl, the protecting group can be removed by reacting with a reducing agent such as the combination of a metal such as zinc with an acid such as acetic acid or hydrochloric acid.

Preferred examples of the reducing agent include the combination of zinc with acetic acid.

There is no particular limitation on the nature of the solvent to be employed, provided that it has no adverse effect on the present reaction. Preferred examples include alcohols such as methanol and ethanol, ethers such as tetrahydrofuran and dioxane, aliphatic acids such as acetic acid and mixtures of said organic solvents and water.

The reaction temperature usually ranges from 0°C to 40 0 C (preferably from 0 C to 30 0 The reaction time depends on the natures of the starting compound and reducing agent; however, it usually ranges from 5 minutes to 12 hours (preferably from minutes to 4 hours).

After the completion of the reaction, a compound is obtained from the reaction mixture by known means. For example, the insoluble matter is filtered off from the reaction mixture, followed by distilling off the solvent, whereby a compound can be obtained.

If necessary, the resulting compound can be purified by known means, for example, by recrystallization, preparative thin-layer chromatography or column chromatography.

On the other hand, Method B is another process for the preparation of compound Described specifically, a compound of formula (VII) is subjected to a reaction with 97162p2.doc P79569/FP-9716(PCTYtsa-igEngish translation/07.10.98 a compound of formula (III) in the presence of a base to give a compound of formula (VI) (Step B1) and then any protecting groups in the compound (IV) are removed to afford a compound (Step B2). The starting compound of formula (VII) used in this synthetic process is prepared by the method disclosed in Japanese Patent Application Kokai No. SHO 62-30781. A description of each step will next be made.

(Step B1) In Step B a compound of formula (VI) is prepared by reacting Compound

(VII)

with a mercaptopyrrolidine derivative (III) in an inert solvent in the presence of a base.

There is no particular limitation on the nature of the solvent to be employed, provided that it has no adverse effect on the present reaction. Examples of suitable solvents include tetrahydrofuran, acetonitrile, dimethylformamide, dimethylsulfoxide and water, and a mixture thereof, of which the acetonitrile is preferred.

There is no particular limitation on the nature of the base to be employed, provided that it does not affect the other part of the compound, particularly the p-lactam ring. Examples of suitable bases include organic bases such as diisopropylethylamine, triethylamine, N-methylpiperidine and 4 -dimethylaminopyridine; and inorganic bases such as potassium carbonate and sodium bicarbonate, of which diisopropylethylamine is preferred.

Although no particular limitation is imposed on the reaction temperature, it is preferred to carry out the reaction at a relatively low temperature in order to suppress side reactions. The reaction temperature usually ranges from -20 0 C to 40 0 C (preferably from -10 0 C to 20 0

C).

The reaction time mainly depends on the reaction temperature or nature of the reaction reagent; however it usually ranges from 15 minutes to 75 hours (preferably from 30 minutes to 18 hours).

After the completion of the reaction, the resulting compound (VI) of this step is obtained from the reaction mixture by known means. To the reaction mixture or a residue available by distilling off the solvent from the reaction mixture, an organic solvent which is not miscible with water is added, followed by washing with water and Doc: 97162p2.doc

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7 9 569/FP-9716(PCxytsa-igEngiish translationiO7.1O.98 distilling off the organic solvent. If necessary, the resulting compound can be further purified by known means; for example, by recrystallization, reprecipitation or chromatography. It is also possible to subject the resulting compound (VI) to the subsequent reaction (step B2) without isolation from the reaction mixture, if necessary.

(Step B2) In the Step B2 a compound is prepared by removal of any protecting groups from the compound (VI).

This step can be accomplished by a similar method to that described in Step A3 of Method A.

The 1-methylcarbapenem compound of the formula thus obtained by Method A or B can be convered into its pharmacologically acceptable salt or derivative (preferably an ester derivative) by a method well known in the field of p-lactam antibiotics.

Incidentally, the mercaptopyrrolidine compound (IV) to be used as a starting material in each of Methods A and B can be prepared by a known method; for example, the methods described in I. Kawamoto et al., Synlett, 575(1995), Japanese Patent Application Kokai No. Hei 2-28180, Japanese Patent Application Kokai No. Hei 2- 3687, Japanese Patent Application No. Hei 4-211083 or Japanese Patent Application Kokai No. Hei 5-339269.

The compounds of formula and pharmacologically acceptable salt thereof of the present invention exhibit strong and well-balanced antibacterial activity against a wide range of bacteria including Gram positive bacteria such as Staphylococcus aureus and Bacillus subtilis, Gram negative bacteria such as Escherichia coli, Shigella species, Klebsiella penumoniae, Proteus species, Serratia species, Enterobacter species and Pseudomonas aeruginosa, and anaerobes such as Bacteroidesfragilis. They also exhibit excellent antibacterial activity against Pseudomonas aeruginosa having resistance against meropenem. In addition, the compounds of the present invention exhibit high stability against p-lactamases and against dehydropeptidase-I, and high Doc: 97162p2.doc

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7 9569/FP-9716(PCT)/tsaig/English translation/07. 10.98 recovery rates in urine. Furthermore, the compounds of the present invention are excellent in in vivo kinetics such as half-life in blood and are relatively free from nephrotoxicity. Judging from these advantages, they are excellent antibiotics.

Compared with aminomethyl compounds [compounds represented by the formula wherein

R

2 and R 3 represent hydrogen atoms; Compound A and Compound B disclosed in Japanese Patent Application Kokai No. Hei 5-310740], the compounds having a substituent at the amino group exhibited superior activity against Pseudomonas aeruginosa, superior in vivo kinetics and lower nephrotoxicity.

Accordingly, the compounds of the formula and pharmacologically acceptable salts or derivatives thereof of the present invention are excellent antibacterial agents for the treatment or prevention (preferably, treatment) of infections caused by various bacteria.

[Capability of Utility in Industry] When compounds or pharmacologically acceptable salts thereof are used as antibacterial agents, they can be administered orally in the form of tablets, capsules, granules, powders or syrups by using them as they are or mixing them with a necessary pharmacologically acceptable additive such as excipient or diluent, or administered parenterally in the form of injections.

The above formulations can be prepared in a known manner by using additives.

Examples of the additives include excipients sugar derivatives such as lactose, sucrose, dextrose, mannitol or sorbitol; starch derivative such as corn starch, potato starch, a-starch, dextrin or carboxymethyl starch; cellulose derivatives such as crystalline cellulose, low-substituted hydroxypropylcellulose, hydroxypropylmethylcellulose, carboxymethylcellulose, carboxymethylcellulose calcium or internally cross-linked carboxymethylcellulose sodium; acacia; dextran; pullulan; silicate derivatives such as light silicic anhydride, synthetic aluminum silicate or magnesium aluminometasilicate; phosphate derivatives such as calcium phosphate; carbonate derivatives such as calcium carbonate; or sulfate derivatives such as calcium sulfate), binders the above-exemplified excipients, gelatin, polyvinyl pyrrolidone; or Macrogol), disintegrators the above-exemplified excipients or chemically P79569/FP-9716(PCT)/tsaig/English translation/07.10.98 modified starch or cellulose derivatives such as cross carmellose sodium, carboxymethyl starch sodium or crosslinked polyvinylpyrrolidone), lubricants talc, stearic acid, metal salts of stearic acid such as calcium stearate or magnesium stearate; colloidal silica; veegum; wax such as spermaceti; boric acid; glycol; carboxylic acids such as fumaric acid or adipic acid; sodium carboxylates such as sodium benzoate; sulfates such as sodium sulfate; leucine; lauryl sulfates such as sodium lauryl sulfate or magnesium lauryl sulfate; silicic acids such as silicic anhydride or silicic hydrate; or starch derivatives exemplified above as the excipient), stabilizers (e.g.

p-hydroxybenzoates such as methyl p-hydroxybenzoate or propyl p-hydroxybenzoate; alcohols such as chlorobutanol, benzyl alcohol or phenylethyl alcohol; benzalkonium chloride; phenol derivatives such as phenol or cresol; thimerosal; acetic anhydride; or sorbic acid), corrigents ordinarily-employed sweeteners, souring agents or flavors), suspending agents Polysorbate 80 or carboxymethylcellulose sodium), diluents and solvents for formulation water, ethanol or glycerin).

The dose of the compounds will vary depending on the condition and age of the patient. Orally, they are administered in an amount of 10 mg (preferably 50 mg) in a single dose as a lower limit and 2000 mg (preferably 1000 mg) in a single dose as an upper limit, while intravenously, they are administered in an amount of 10 mg (preferably 100 mg) in a single dose as a lower limit and 3000 mg (preferably 2000 mg) in a single dose as an upper limit. It is desirable to be administered to an adult in a single dose or in divided dose (sixth) per day depending on the condition of the patient.

[Best Modes for Carrying out the Invention] The present invention will hereinafter be described in more detail by examples, referential examples, tests and formulation examples. However the present invention is not limited to or by these examples. Incidentally, in the nuclear magnetic resonance spectrum in the examples and referential examples, sodium trimethylsilylpropionate-d 4 was used as an internal standard for the measurement in heavy water, while tetramethylsilane was used as an internal standard in the other solvents, unless otherwise indicated.

P

7 9569/FP-9716(PCr)/tsaigEnglish translation/07.1 0.98 (iExample 1) (I R, 5 S, 6

S)-

2 2 S, 4 R)-3 -Am inom ethylpyrrol idin- 1 -ylcarbonyl]- 1 methylpyrrolidin-4-ylthio]-6-r(I1R)- 1 -hydroxyethyl]- I1-methyl- I -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No, 61) OH CH3 H

ICNNH

CH

3 IS

N

N

CH

3 0

COOH

To a solution of 4-nitrobenzyl (1R, 5R, 1R)-l1-hydroxyethyl]- 1methyl-2-(diphenylphosphoryloxy) 1 -carbapen-2-em-3-carboxylate (0.72 g) in anhydrous acetonitrile (7 ml), NN-diisopropylethylamine (0.21 ml) and a solution of (2S,4 S)-4-mercapto-l1-methyl-2-[(3R)-3 4 -nitrobenzyloxycarbonylaminomethyl)pyrrolidin-1-ylcarbonyl]pyrrolidine (0.54 g) in anhydrous acetonitrile (10 ml) were added while stirring in an ice bath. The resulting mixture was stirred overnight at 0 0

C.

To the reaction mixture, ethyl acetate was added. The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column ethyl acetate/methanol whereby 4nitrobenzyl (1R, 5 S, 1R)-l1-hydroxyethyl]-l1-methyl-2-[(2S,4S)-l1-methyl-2- 4 -nitrobenzyloxycarbonylaminomethyl)pyrrolidin-1 -ylcarbonyl]pyrrolidin-4.

ylthio]- 1 -carbapen-2-em-3 -carboxyl ate (0.74 g) was obtained.

Infrared absorption spectrum (KBr) v max m1 1771,1721,1637,1608,1522,1490,1274,1245,1210,1181,1137,1107,1074,1046,1026, 1014.

Nuclear magnetic resonance spectrum (270 IvLHz, CDCI 3 8 PPM: 1 .27(3H,d,J=7.3H 2 1 .36(3Hd,J=6.6Hz), 1.45-2. 8O(5HKm), 2.31,2.3 1(3H,sx2), 2.98- 3.90(1 3H,m), 4.1 S- 4 3 0(21-Im), 5.10-5. 50(4H,m), 7.48(2H,d,J=8.6Hz), 7.63(2H,d,J=8.6Hz), 8.20(4Hd,J=8.6Hz).

To a solution of the compound (0.73 which had been obtained in in tetrahydrofuran (40 ml) and water (28 ml), a 10% palladium-carbon catalyst (1.46 g) Doc: 97162p2.doe Doc: 7162p.docP 7 9569/FP-9716(PCT)/ts-ig/English taslationiO7. 10.98 was added. The resulting mixture was allowed to absorb hydrogen for 1.5 hours while stirring at an external temperature of 30 0 C. The catalyst was then filtered off. The filtrate was washed with ether and ethyl acetate, followed by concentration by evaporation under reduced pressure. The residue was separated by reversed-phase column chromatography ["Cosmosil 75C18-PREP" (NACALAI TESQUE,

INC.),

acetonitrile/water 6/94 1/9 (elution while an acetonitrile concentration was increased gradually from 6% to followed by concentration by evaporation under reduced pressure and lyophilization, whereby crude title compound (246 mg) was obtained as a powder.

Out of the 246 mg of the crude title compound, a 230 mg portion was separated and purified through an HPLC preparative column ["Cosmosil 5C18-AR"

(NACALAI

TESQUE, INC.), acetonitrile/water 6/94 1/9 (elution while an acetonitrile concentration was increased gradually from 6% to followed by concentration by evaporation under reduced pressure and lyophilization, whereby the title compound (156 mg) was obtained as a powder.

Infrared absorption spectrum (KBr) v max cm1: 1758,1633,1594,1488,1455,13851385,1340,1313,1253,1225,1210,1181,1149,1095.

Nuclear magnetic resonance spectrum (400 MHz, D 2 0) 6 ppm: 1.21(3H,d,J=7.2Hz), 1.30(3H,d,J=6.4Hz), 1.63-1.86(2H,m), 2.14-2.93(4H,m), 2.32(3H,s), 3 03 -4.01(13H,m), 4.17-4.30(2H,m).

(Example 2) 3 -Aminomethylpyrrolidin- -ylcarbonyl]-lmethylpyrrolidin-4-ylthio-6-[(1R)-l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3carboxylic acid (Stereoisomer of Exemplified Compound No. 61) H H 3

CONNH

I

COOH

P79569/FP-976(PCTytta.igEnglish translation/07. 10.98 To a solution of 4-nitrobenzyl (1R,5R,6S)-6-[(1R)-1-hydroxyethyl]-1methyl-2-(diphenylphosphoryloxy)-l-carbapen-2-em-3-carboxylate (0.73 g) in anhydrous acetonitrile (7 ml), N,N-diisopropylethylamine (0.22 ml) and a solution of (2S,4S)-4-mercapto-1-methyl-2-[(3S)-3-( 4 -nitrobenzyloxycarbonylaminomethyl)pyrrolidin-1-ylcarbonyl]pyrrolidine (0.57 g) in anhydrous acetonitrile (10 ml) were added while stirring in an ice bath. The resulting mixture was stirred overnight at 0°C.

To the reaction mixture, ethyl acetate was added. The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol whereby 4nitrobenzyl (1R,5S,6S)-6-[(1R)-1-hydroxyethyl]-l-methyl-2-[(2S,4S)-1-methyl-2-[(3S)- 3 4 -nitrobenzyloxycarbonylaminomethyl)pyrrolidin- -ylcarbonyl]pyrrolidin-4-ylthio]- 1-carbapen-2-em-3-carboxylate (0.78 g) was obtained.

Infrared absorption spectrum (KBr) v max cm' 1 1771,1721,1638,1608,1522,1490,1453,1375,1347,1324,1274,1246,1210,1181,1137.

Nuclear magnetic resonance spectrum (270 MHz, CDC13) 6 ppm: 1.25(3H,d,J=7.3Hz), 1.36(3H,d,J=6.6Hz), 1.46-2.80(5H,m), 2.31(3H,s), 2.98- 4.00(13H,m), 4.18-4.31(2H,m), 5.11-5.50(4H,m), 7.42-7.67(4H,m), 8.13-8.25(4H,m).

To a solution of the compound (0.76 which had been obtained in in tetrahydrofuran (40 ml) and water (28 ml), a 10% palladium-carbon catalyst (1.55 g) was added. The resulting mixture was allowed to absorb hydrogen for 1.5 hours while stirring at an external temperature of30 0 C. The catalyst was then filtered off. The filtrate was washed with ether and ethyl acetate, followed by concentration by evaporation under reduced pressure. The residue was separated by reversed-phase column chromatography ["Cosmosil 75C18-PREP" (NACALAI TESQUE, INC.), acetonitrile/water 6/94 1/9 followed by concentration by evaporation under reduced pressure and lyophilization, whereby crude title compound (248 mg) was obtained as a powder. Out of 248 mg of the compound, a 189 mg portion was separated and purified through an HPLC preparative column ["Cosmosil 5C18-AR" (NACALAI TESQUE, INC.), acetonitrile/water 6/94 1/9 followed by concentration by Doc: 97162p2.doc

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7 9 5 69/FP-9716(PCT)/tsa.ig/English translation/07.10.98 evaporation under reduced pressure and lyophilization, whereby the title compound (125 mg) was obtained as a powder.

Infrared absorption spectrum (KBr) v maxcm: 1759, 1633, 1595, 1488, 145 5,13 84, 1253, 1211, 1180, 1148, 1095.

Nuclear magnetic resonance spectrum (400 MfHz, D 2 0) 5 PPM: 1.2 1(3H,d,J=7.2Hz), 1 .30(3H,d,J=6.4Hz), 1.63-1 .87(1H~m),2. l3-2.4O(1H~m), 2.31 ,2.30(3H,sx2), 2.53-2.93(3H,m), 3.08-3.90(1 1H,m), 4.1 8 -4.29(2H~m).

(Example 3) (1 R, 5S,6S)-6-r( 1R)-l1-Hydroxyethyl]-l1-methyl-2-[(2 S,4 S-2-r(3R)-3 m ethyl am inomethylpyrrolidin- 1 -ylcarbonyllpyrrolidin-4-ylthiol- 1 -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No. 1) OH H H, HH H H ICON\

NCH

OH

3 I-

NN

O

COCH

To a solution of 4-nitrobenzyl (iR.,5R,6S)-6-[(1R)-l1-hydroxyethyl]- 1methyl-2-(diphenylphosphoryloxy) 1 -carbapen-2-em-3 -carboxylate (1.76 g) in anhydrous acetonitrile (20 ml), NN-diisopropylethylamine (0.51 ml) and a solution of (2S,4S)-4-mercapto-2-[(3 S)-3 -(N-methyl-N-4-nitrobenzyloxycarbonylaminomethyl)pyrrolidin-1I-ylcarbonyl]- 1-( 4 -nitrobenzyloxycarbonyl)pyrrolidine (1.92 g) in anhydrous acetonitrile (20 ml) were added while stirring in an ice bath. The resulting mixture was stirred overnight at 0 0 C. To the reaction mixture, ethyl acetate was added. The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure.

The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol 95/5 whereby 4-nitrobenzyl (1R,5S,6S)-6-[(1R)-lhydroxyethyl]-l1-methyl-2-[(2S,4S)-2-[(3 S)-3 -(N-methyl-N-4-nitrobenzyloxycarbonylaminomethyly. 1-( 4 -nitrobenzyloxycarbonyl)pyrrolidin 1 -ylcarbonyl}..

pyrrolidin-4-ylthio])-1-carbapen-2-em-3 -carboxylate (2.52 g) was obtained.

Doc: 97162p2.doe Doc: 7162p.docP 7 9569IFP-97l6(PCTytsa-ig/English translationO7. 10.98 Infrared absorption spectrum v max m1 1774, 1707, 1652, 1607, 1522, 144 1, 1404, 1346, 1295, 12 10, 1143, 1110.

Nuclear magnetic resonance spectrum (400 MlHz, CDCI 3 5 PPM: 1 .29(3H,d,J=7.2Hz), 1 .37(3H,d,J=6.1Hz), 1 .55-2.73(5HKm), 2.93-4.58(1 8H,m), 5.02- 5.52(6H,m), 7.40-7.70(6H~m), 8.lS-8.30(6H,m).

To a solution of the compound (2.52 which had been obtained in in tetrahydrofuran (120 ml) and water (84 ml), a 10% palladium-carbon catalyst (5.07 g) was added. The resulting mixture was allowed to absorb hydrogen for 1.5 hours while stirring at an external temperature of 30'C. The catalyst was then filtered off. The filtrate was washed with ether and ethyl acetate, followed by concentration by evaporation under reduced pressure. The residue was purified by reversed-phase column chromatography ["Cosmosil 75C18-PREP" (NACALAI TESQUE, INC.), acetonitrile/water 6/94 8/92 followed by concentration by evaporation under reduced pressure and lyophilization, whereby the title compound (406 mg) was obtained as a powder.

Infrared absorption spectrum (KBr) v max cm-1: 1757, 1634, 1598, 1456, 1386, 13 11, 1284, 1257, 1225, 1180, 1148, 1099.

Nuclear magnetic resonance spectrum (400 MiHz, D 2 0) 5 PPM: 1 .22(3H,d,J=7.2Hz), 1 .30(3H,d,J=6.4Hz), l.58-1.87(2HKm), 2.16-2.3 1(1H,m), 2.58- 2. 80(2H,m), 2.76(3H,s), 3.03-4. 07(12H,m), 4.1 8-4.29(2HKm).

(Example 4) (1 SS,6S 1R)-l1-Hydroxyethyl]-l1-methyl-2-[(2S,4S)-2-[(3R)-3 methylaminomethylpyrrolidin-1 -ylcarbonyll-.1 -methylpyrrolidin-4-ylthio]-lI-carbapen- 2-em-3-carboxylic acid (Stereoisomer of Exemplified Compound No. 62) H OHHCH 3 CON(D NH H NH N\ 3 Doc: 97162p2.doc Doc: 7162p.docP 7 9 5 69/FP-9716(PCT)zsa-ig/English translationl07. 10.98 To a solution of 4-nitrobenzyl (1 R, 5R,6S)6-)-6-[(1R)--yohydroxyethyl]methyl-2-(diphenylphosphoryloxy)-l-carbapen-2-em-3-carboxylate (0.72 g) in anhydrous acetonitrile (5 ml), N,N-diisopropylethylamine (0.21 ml) and a solution of (2S,4S)-4-mercapto-l-methyl-2-[(3 S)-3-(N-methyl-N-4nitrobenzyloxycarbonylaminomethyl)pyrrolidin--ylcarbonyl]pyrrolidine (0.54 g) in anhydrous acetonitrile (5 ml) were added while stirring in an ice bath. The resulting mixture was allowed to react overnight at 0°C. To the reaction mixture, ethyl acetate was added. The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol whereby 4-nitrobenzyl (1R,5S,6S)-6-[(1R)-1hydroxyethyl]-1 -methyl-2-[(2S,4S)-1 -methyl-2-[(3 S)-3-(N-methyl-N-4nitrobenzyloxycarbonylaminomethyl)pyrrolidin-1 -ylcarbonyl]pyrrolidin-4-ylthio]- 1 carbapen-2-em-3-carboxylate (0.46 g) was obtained.

Infrared absorption spectrum (KBr) v max cm- 1 1770,1705,1640,1607,1522,1489,1452,1403,1376,1346,1278,1210,1143,1106.

Nuclear magnetic resonance spectrum (400 MHz, CDCl 3 6 ppm: 1.27(3H,d,J=7.3Hz), 1.37(3H,d,J=6.2Hz), 1.49-2.83(5H,m), 2.31,2.35(3H,s2), 2.95- 3.80(13H,m), 4.20-4.30(2H,m), 5.16-5.53(4H,m), 7.52(2H,d,J=8.8Hz), 7.64,7.65(2H,dx 2,J=8.8Hz), 8.22,8.23(4H,dx2,J=8.8Hz).

To a solution of the compound (0.46 which had been obtained in in tetrahydrofuran (22 ml) and water (15 ml), a 10% palladium-carbon catalyst (0.93 g) was added. The resulting mixture was allowed to absorb hydrogen for 1.5 hours while stirring at an external temperature of 30 0 C. The catalyst was then filtered off. The filtrate was washed with ether and ethyl acetate, followed by concentration by evaporation under reduced pressure. The residue was purified by reversed-phase column chromatography ["Cosmosil 75C18-PREP" (NACALAI TESQUE, INC.), acetonitrile/water 4/96 8/92 followed by concentration by evaporation under reduced pressure and lyophilization, whereby the title compound (150 mg) was obtained as a powder.

Doc: 97162p2.doc P79569IFP-97 6(PCy)/tsa-ig/English translationl07.1 0.98 Infrared absorption spectrum (KBr) v maxcm: 1760, 1634, 1598, 1486,1454,1383,13 14,1279,1253,1221,1180,1148,1095.

Nuclear magnetic resonance spectrum (400 M~z, D 2 0) 5 PPM: 1 .21(3H,d,J=7.21{z), 1 .30(3H,dj=6.2Hz), l.

6 5 -l.86(2HKm), 2.1 5-2.30(1H,m), 2.38(3H,s), 2.57-3.00(2H,m), 3.10-3 .99(12H,m), 4.1 8-4.29(2HKm).

(Example (iR 5S,6S)-6-r( IR)-l1-Hydroxyethyl]-l1-methyl-2-[(2S,4S)-2-r(3 S)-3methylaminomethylpyrrolidin- 1 -ylcarbonyllpyrrolidin-4-ylthio]. 1 -carbapen-2-em-3carboxylic acid (Stereoisomer of Exemplified Compound No. 1) O HH

CH

3 H CON

NHCH

3

CH

3 z

S

NN

0

COOH

To a solution of 4-nitrobenzyl (11R, 5R, 1R)-l1-hydroxyethyl]- 1methyl-2-(diphenylphosphoryloxy) 1 -carbapen-2-em-3 -carboxylate (0.85 g) in anhydrous acetonitrile (5 ml), NN-diisopropylethylamine (0.25 ml) and a solution of (2 S,4 S)-4-mercapto-2- R)-3 ethyl-N-4-nitrobenzyloxycarbonylam inom ethyl)pyrrolidin- 1-ylcarbonyl]- 1-( 4 -nitrobenzyloxycarbonyl)pyrrolidine (0.90 g) in anhydrous acetonitrile (10 ml) were added while stirring in an ice bath. The resulting mixture was stirred overnight at 0 0 C. To the reaction mixture, ethyl acetate was added. The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure.

The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol 95/5 whereby 4-nitrobenzyl (1R,5S,6S)-6-[(1R)-ilhydroxyethyl]- 1 -m ethylI-2-[(2 S,4 -(N-methyl-N-4nitrobenzyloxycarbonylaminomethyl). 1 4 -nitrobenzyloxycarbonyl)pyrrolidin 1ylcarbonyl]pyrrolidin-4-ylthio]-l1-carbapen-2-em-3 -carboxylate (1.07 g) was obtained.

Doe: 97162p2.doe Doc: 7162p.docP795691FP-9716(PCT)/tsa-ig/English translationO7.1O.98 Infrared absorption spectrum (KBr) v max m1 1773,1707,1654,1607,1522,1441,1404,13 73,1346,1295,1210,1180,1143,1110,1047, 1015.

Nuclear magnetic resonance spectrum (400 MHz, CDCl 3 8 PPM: 1 .27(3H,d,J=7.3Hz), 1 .37(3H,d,J=6.3Hz), 1 .50-2.72(SHKm), 2.90-4.3 1(14H,m), 4.40- 4.55(1H,m), 5.OI-5.52(6K{m), 7.41-7.67(6H,m), 8. l3-8.27(6H,m).

To a solution of the compound (1.05 which had been obtained in in tetrahydrofuran (50 ml) and water (35 ml), a 10% palladium-carbon catalyst 10 g) was added. The resulting mixture was allowed to absorb hydrogen for 1.5 hours while sturring at an external temperature of 30 0 C. The catalyst was then filtered off. The filtrate was washed with ether and ethyl acetate, followed by concentration by evaporation under reduced pressure. The residue was purified by reversed-phase column chromatography ["Cosmosil 75C 18-PREP" (NACALAI TESQUE, INC.), acetonitrile/water =6/94 followed by concentration by evaporation under reduced pressure and lyophilization, whereby the title compound (127 mg) was obtained as a powder.

Infrared absorption spectrum (KLBr) v max m1 1756, 1633, 1598, 1457, 13 87, 13 11, 1285, 1258, 1226, 118 1,1149, 1096, 1074.

Nuclear magnetic resonance spectrum (400 MHz, D 2 0) 5 PPM: 1 .22(3H,d,J=7.2FHz), 1 .30(3H,d,J=6.4Hz), 1.59-1 .87(2HKm), 2. 16-2.32(1H,m), 2.58- 2.8 1(2H,m), 2.77(3H,s), 3 .05-3.86(12H,m), 3 .94-4.03(1H~m), 4.1 8-4.28(2H,m).

(Example 6) (1 R,5 S,6S)-6-r( 1R)-l1-Hydroxyethyl.. 1-methyl-2-[(2S,4S)-2-[(3 S)-3methylaminomethylpyrrolidin -ylcarbonyl]- I-methylpyrrolidin-4-ylthio.. 1-carbapen- 2-em-3-carboxylic acid (Stereoisomer of Exemplified Compound No. 62) OH CH 3

HH

H H

CNHH

3

CH

3 I

T

N

OH

3 Doc: 97162p2.doc Doc: 7162p.docP 7 9 5 6 9/FP-9716(PCT)ftsa-igEnglish translatior,107 10.98 To a solution of 4-nitrobenzyl -hydroxyethyl]-1methyl-2-(diphenylphosphoryloxy)- 1-carbapen-2-em-3-carboxylate (0.66 g) in anhydrous acetonitrile (5 ml), N,N-diisopropylethylamine (0.19 ml) and a solution of (2S,4S)-4-mercapto-1 -methyl-2-[(3R)-3-(N-methyl-N-4nitrobenzyloxycarbonylaminomethyl)pyrrolidin- 1-ylcarbonyl]pyrrolidine (0.53 g) in anhydrous acetonitrile (5 ml) were added while stirring in an ice bath. The resulting mixture was allowed to react overnight at 0°C. To the reaction mixture, ethyl acetate was added. The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column ethyl acetate/methanol whereby 4-nitrobenzyl (1R,5S,6S)-6-[(1R)-1hydroxyethyl]-l-methyl-2-[(2S,4S)-1-methyl-2-[(3R)-3-(N-methyl-N-4nitrobenzyloxycarbonylaminomethyl)pyrrolidin-1-ylcarbonyl]pyrrolidin-4-ylthio]-1carbapen-2-em-3-carboxylate (0.43 g) was obtained.

Infrared absorption spectrum (KBr) v max cm'': 1771,1734,1705,1639,1608,1522,1489,1452,1403,1375,1346,1278,1246,1210,1142.

Nuclear magnetic resonance spectrum (400 MHz, CDCl 3 6 ppm: 1.27(3H,d,J=7.2Hz), 1.37(3H,d,J=6.2Hz), 1.52-2.78(8H,m), 2.94-3.82(13H,m), 4.19- 4.30(2H,m), 5.12-5.50(4H,m), 7.50-7.68(4H,m), 8.21-8.23(4H,m).

To a solution of the compound (0.42 which had been obtained in in tetrahydrofuran (20 ml) and water (15 ml), a 10% palladium-carbon catalyst (0.83 g) was added. The resulting mixture was allowed to absorb hydrogen for 1.5 hours while stirring at an external temperature of 30 0 C. The catalyst was then filtered off. The filtrate was washed with ether and ethyl acetate, followed by concentration by evaporation under reduced pressure. The residue was purified by reversed-phase column chromatography ["Cosmosil 75C18-PREP" (NACALAI TESQUE, INC.), acetonitrile/water 6/94 8/92], followed by concentration by evaporation under reduced pressure and lyophilization, whereby the title compound (28 mg) was obtained as a powder.

Infrared absorption spectrum (KBr) v max cm' 1 1757,1642,1597,1489,1457,1384,1251,1210,1158,1095,1028.

Doc: 97162p2.doc

P

7 9 56 9 IFP-9716(PCT)Ttsai.giEnglish translation07. 10.98 Nuclear magnetic resonance spectrum (400 MlHz, D 2 0) 5 PPM: 1l.21(3H,d,J=7.2Hz), 1 .30(3H,d,J=641z), 1.67-1 .88(2HKm), 2.1 7-2.32(1H,m), 1(3H,br.s), 2.57-2.75(1H,m), 2.76(3H,s), 2.87-3.01(1H,m), 3.05-3.87(1 1H~m), 3.90- 4.00(1H,m), 4. 18-4.29(2HKm).

(Example 7) 5 S, 6

S)-

2 2

S,

4 S)-2-r(3R)-3-Guanidinomethylpyrrolidin-1 -ylcarbonyllpyrrolidin-4ylthio] 1 -hydroxyethyl]- 1 -m ethyl- I -carb apen-2-em-3 -carboxylic acid (Stereoisomer of Exemplified Compound No. 18) OH H H H HH H NH CO

NH

2

OH

3 C3NH

NH

0

COOH

To a solution of 4-nitrobenzyl (1 R, 1R)-l1-hydroxyethyl]-1methyl-2-(diphenylphosphoryloxy> 1 -carbapen-2-em-3 -carboxylate (0.70 g) ill anhydrous acetonitrile (10 ml), N,N-diisopropylethylamine (0.21 ml) and a solution of (2 S,4S)-4-mercapto- l-( 4 -nitrobenzyloxycarbonyl)-2[(3R)3(4-nitrobenzyloxycarbonylguanidinomethyl)pyrrolidin-.1 -ylcarbonyl]pyrrolidine (1.28 g) in anhydrous acetonitrile (10 ml) were added while stirring in an ice bath. The resulting mixture was allowed to react overnight at 0 0 C. To the reaction mixture, ethyl acetate was added.

The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure.

The residue was purified by chromatography through a silica gel column (ethyl acetate/m ethanol 95/5 9/1 whereby 4-nitrobenzyl (1R,5S,6S)-6-[(1R)-1hydroxyethyl]-lI-methyl-2-[(2S,4S)- I-(4-nitrobenzyloxycarbonyl)-2-[(3R)3(4nitrobenzyloxycarbonylguanidinomethyl)pyrrolidin-1 -ylcarbonyl]pyrrolidin-4-ylthio}..

l-carbapen-2-em-3-carboxylate (0.97 g) was obtained.

Infrared absorption spectrum (KBr) v maxcm: 1772, 1709, 1647, 1608,1522, 1491, 1440, 1404, 1378, 1346, 1322, 1287, 1210, 1178,1133, 1109.

Doc: 97162p2.doc Doc: 7162p.docP 7 9569/FP-9716(PCT)/tsa-ig/English translationO7. 10.98 Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 8 ppm: 1.28(3H,d,J=7.4Hz), 1.37(3H,d,J=6.3Hz), 1.50-2.77(8H,m), 2.94-4.05(12H,m), 4.18- 4.28(2H,m), 4.40-4.55(1H,m), 5.10-5.51(6H,m), 7.37-7.68(6H,m), 8.10- 8 2 7 6 H,m).

To a solution of the compound (0.95 which had been obtained in in tetrahydrofuran (50 ml) and water (35 ml), a 10% palladium-carbon catalyst (1.90 g) was added. The resulting mixture was allowed to absorb hydrogen for 1.5 hours while stirring at an external temperature of 30 0 C. The catalyst was then filtered off. The filtrate was washed with ether and ethyl acetate, followed by concentration by evaporation under reduced pressure. The residue was purified by reversed-phase column chromatography ["Cosmosil 75C18-PREP" (NACALAI TESQUE, INC.), acetonitrile/water 8/92 12/88], followed by concentration by evaporation under reduced pressure and lyophilization, whereby crude title compound (224 mg) was obtained as a powder.

The crude title compound was purified through an HPLC preparative column ["Cosmosil 5C18-AR" (NACALAI TESQUE, INC.), acetonitrile/water 8/92 12/88], followed by concentration by evaporation under reduced pressure and lyophilization, whereby the title compound (166 mg) was obtained as a powder.

Infrared absorption spectrum (KBr) v max cm-: 1757,1630,1455,1386,1313,1283,1260,1224,1182,1147,1102,1074.

Nuclear magnetic resonance spectrum (400 MHz, D 2 0) 8 ppm: 1.22(3H,d,J=7.2Hz), 1.30(3H,d,J=6.2Hz), 1.55-1.84(2H,m), 2.18-2.23(1H,m), 2.52- 2.78(2H,m), 3.06(1H,dd,J=12.3,3.6Hz), 3.13-3.85(10H,m), 3.96-4.03(1H,m), 4.18- 4.29(2H,m).

Doc: 97162p2.doc P79569/F-9716(PCT),tsa-ig/English translation/07.10.98 (Example 8) (1 R,5 S, 6

S)-

2 2

S,

4

S)-

2 3 R)-3-Acetimidoylaminomethylpyr-rolidinylcarbonyl~lpyrrolidin-4-ylthio] 1R)-l1-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No. 17) OH H H H H 7Hi C C N

H

3

S

OH

3 -K N NH NH O

COOH

To a solution of 4-nitrobenzyl (1R, 5R, 1R)-l1-hydroxyethyl]- 1methyl-2-(diphenylphosphoryloxy). 1 -carb apen-2-em-3 -carboxyl ate (0.81 g) in anhydrous acetonitrile (10 ml), NN-diisopropylethylamine (0.44 ml) and a solution of (2S,4S)-4-mercapto- 1 4 -nitrobenzyloxycarbonyl)-2[(3R)3(4-nitrobenzyloxycarbonylacetim idoylaminomethyl)pyn-olidin- 1 -ylcarbonyl] pyrroli dine (1.35 g) in anhydrous acetonitrile (15 ml) were added while stirring in an ice bath. The resulting mixture was allowed to react overnight at 0 0 C. To the reaction mixture, ethyl acetate was added. The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol 95/5 9/1 whereby 4-nitrobenzyl (1R,,5S,6S)- 1R)-l1-hydroxyethyl]-l1-methyl-2-[(2S,4 1-(4-nitrobenzyloxycarbonyl)-2-[(3R)-3 4 -nitrobenzyloxycarbonylacetimidoaminomethyl)pyrrolidin- 1 -ylcarbonyl]pyrrolidin-4ylthio]- 1 -carbapen-2-em-3-carboxylate (1.15 g) was obtained.

Infrared absorption spectrum (KBr) v max cm- 1 1773,1709,1650,1607,1556,1522,1494,1441,1404,13 73,1246,1278,123 7,1212,1126, 1110.

Nuclear magnetic resonance spectrum (400 MvHz, CDCl 3 8 PPM: 1 .29(3H,d,J=7. 1Hz), 1 .37(3H,d,J=6. 1Hz), 1 .50-2.72(9HKm), 2.98-4.06(12HKm), 4.22- 4.30(2H,m), 4.46-4. 57(l1Hm), 5.1 6-5.52(6H,m), 7.43-7.67(6H,m), 8.1 7-8.27(6HKm).

Doc: 97162p2.doc Doc: 7162p.docP79569fFP-9716(PCT)tsa-ig/English translation/07. 10.98 To a solution of the compound (1.13 which had been obtained in in tetrahydrofuran (60 ml) and water (42 ml), a 10% palladium-carbon catalyst (2.28 g) was added. The resulting mixture was allowed to absorb hydrogen for 1.5 hours while stirring at an external temperature of30 0 C. The catalyst was then filtered off. The filtrate was washed with ether and ethyl acetate, followed by concentration by evaporation under reduced pressure. The residue was purified by reversed-phase column chromatography ["Cosmosil 75C18-PREP" (NACALAI TESQUE, INC.), acetonitrile/water 8/92 12/88], followed by concentration by evaporation under reduced pressure and lyophilization, whereby a compound (337 mg) was obtained as a powder.

The resulting compound was purified through an HPLC preparative column ["Cosmosil 5C1I 8-AR" (NACALAI TESQUE, INC.), acetonitrile/water 8/92 12/88], followed by concentration by evaporation under reduced pressure and lyophilization, whereby the title compound (254 mg) was obtained as a powder.

Infrared absorption spectrum (KBr) v max cm-1 1756,1684,1633,1593,1455,1386,1312,1284,1261,1226,1182,1284,1261,1226,1182, 1148.

Nuclear magnetic resonance spectrum (400 M1Hz, D 2 0) 5 ppm: 1.22(3H,d,J=7.2Hz), 1.30(3H,d,J=6.4Hz), 1.56-1.67(1Hm), 1.69-1.87(1H,m), 2.12- 2.26(1H,m), 2.25(3H,s), 2.58-2.78(2H,m), 3.06(1Hdd,J=12.3,3.5Hz), 3.14- 3.85(10OHm), 3.96-4.04(IHm), 4.18-4.28(2H,m).

(Example 9) 6

S)-

2 2 S,4S)-2-[(3R)-3-Cyclopropylaminomethylpyrrolidin1ylcarbonyl]pyrrolidin-4-ylthio]-6-(l 1-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3carboxylic acid (Stereoisomer of Exemplified Compound No.

COOH

Doc: 97162p2.doc P79569/FP-9716(PCT)Ia-i/Engish translationl07. 10.98 To a solution of 4 -nitrobenzyl (IR,5R,6S)-6-[(IR)-1-hydroxyethyl]-1methyl-2-(diphenylphosphoryloxy)-1 -carbapen-2-em-3-carboxylate (0.66 g) in anhydrous acetonitrile (7 ml), N,N-diisopropylethylamine (0.19 ml) and a solution of (2S,4S)-4-mercapto- 1 4 -nitrobenzyloxycarbonyl)-2-[(3 S)-3-(N-cyclopropyl-N-4nitrobenzyloxycarbonylaminomethyl)pyrrolidin- 1 -ylcarbonyl]pyrrolidine (0.14 g) in anhydrous acetonitrile (8 mi) were added while stirring in an ice bath. The resulting mixture was allowed to react overnight at 0 0 C. To the reaction mixture, ethyl acetate was added. The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol 95/5), whereby 4-nitrobenzyl (1R,5S,6S)-6-[(1R)-1hydroxyethyl]- 1 -methyl-2-[(2S,4S)-1 4 -nitrobenzyloxycarbonyl)-2-[(3S)-3-(Ncyclopropyl-N-4-nitrobenzyloxycarbonylami 1nomethyl)pyrrolidinylcarbonyl]pyrrolidin-4-ylthio]-1-carbapen-2-em-3-carboxylate (0.80 g) was obtained.

Infrared absorption spectrum (KBr) v max cm-1: 1774,1708,1652,1607,1522,1496,1444,1404,1346,1287,1210,1181,1138 1110.

Nuclear magnetic resonance spectrum (400 MHz, CDCI 3 6 ppm: 0.60- 0.92(4H,m), 1.29(3H,dxJ=7.4Hz), 1.37,1.38(3H,dx2,J=6.3Hz), 1.48-2.74(6Hm), 3.00- 4.05(11H,m), 4.22-4.30(2H,m), 4.36-4.56(1H,m), 5.16-5.52(6H,m), 7.43-7.68(6Hm), 8.17-8.26(6H,m).

To a solution of the compound (0.78 which had been obtained in in tetrahydrofuran (40 ml) and water (28 ml), a 10% palladium-carbon catalyst (1.57 g) was added. The resulting mixture was allowed to absorb hydrogen for 1.5 hours while stirring at an external temperature of 30 0 C. The catalyst was then filtered off. The filtrate was washed with ether and ethyl acetate, followed by concentration by evaporation under reduced pressure. The residue was purified by reversed-phase column chromatography ["Cosmosil 75C18-PREP" (NACALAI TESQUE,

INC.),

acetonitrile/water 6/94 1/9 followed by concentration by evaporation under reduced pressure and lyophilization, whereby the title compound (106 mg) was obtained as a powder.

Doc: 97162p2.doc P79569/FP-9716(PCT)tsa-igFingih translationl07. 10.98 infrared absorption spectrum (KBr) v max cm- 1 1759, 163 7, 1599, 145 5, 1386, 13 12, 1283,1259,1224,1180,1147,1103.

Nuclear magnetic resonance spectrum (400 MHz, D 2 0) 5 PPM: 0.73- O.88(4H,m), 1 .22(3H,d,J=7.2Hz), 1.30(3HKd,J=6.4Hz), 1.63-1 .86(2H,m), 2.14- 2.30(1H,m), 2.58-2.83(3HKm), 3.08-3.92(1 1H,m), 4.06-4. 14(1H,m), 4. 19-4.30(2HKm).

(Example (1 R, 5S,6 S)-2-7(2 S,4 S)-2-r(3 S)-3 -Acetimidoylaminomethylpyrrolidinyl carbonyllpyrroli din-4-ylthio 1 1 -hydroxyethyl]-I 1-m ethyl- I -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No. 17) OH H 3 HH

H

He,1 H H CH3CoN N CH 3

CH

3 N I NH O

COOH

To a solution of 4-nitrobenzyl (1 R, 5R, 1R)-l1-hydroxyethyl]- 1methyl-2-(diphenylphosphoryloxy)- 1 -carbapen-2-em-3 -carboxyl ate (0.58 g) in anhydrous acetonitrile (10 ml), NN-diisopropylethylamine 17 ml) and a solution of (2S,4S)-4-mercapto-l1-( 4 -nitrobenzyloxycarbonyl)-2-[(3 S)-3 4 -nitrobenzyloxycarbonylacetimidoylaminomethyl)pyrrolidin-1 -ylcarbonyl]pyrrolidine (0.93 g) in anhydrous acetonitrile (10 ml) were added while stirring in an ice bath. The resulting mixture was stirred overnight at 0 0 C. To the reaction mixture, ethyl acetate was added.

The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure.

The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol whereby 4-nitrobenzyl (1 R, 5S, 1R)-l1-hydroxyethyl]- 1methyl-2-[(2S,4S)- 1-( 4 -nitrobenzyloxycarbonyl)-2-[(3

S)-

3 -(4-nitrobenzyloxycarbonylacetimidoylaminomethyl)pyrrolidin- 1 -ylcarbonyl]pyrrolidin-4-ylthio]- 1carbapen-2-em-3 -carboxyl ate (0.61 g) was obtained.

Doc: 97162p2.doc Doc:9712p2.docP 7 9 56 9/FP-9716(PCT)/tsa.ig/English translationO7. 10.98 Infrared absorption spectrum (KBr) v max m1 1774,1709,1651,1607,1558,1522,14961441,1404,1373,13461278,1238,1212,1126, 1110,1074,1015.

Nuclear magnetic resonance spectrum (400 MIHz, CDCl 3 8 PPM: 1 .29(3H,d,J=7. 1Hz), 1 .37(3H,d,J=6.2Hz), 1 .55-2.75(9HKm), 3.05-4.06(1 1H,m), 4.22- 4.27(2H,m), 4.44-4.60(1H~m), 5. 17-5.53(6H,m), 7.42-7.68(6H,m), 8.1 5-8.26(6HKm).

To a solution of the compound (0.58 which had been obtained in in tetrahydrofuran (3 0 ml) and water (20 ml), a 10% palladium-carbon catalyst (1.15 g) was added. The resulting mixture was allowed to absorb hydrogen for 1.5 hours while stirring at an external temperature of 30'C. The catalyst was then filtered off. The filtrate was washed with ether and ethyl acetate, followed by concentration by evaporation under reduced pressure. The residue was purified by reversed-phase column chromatography ["Cosmosil 75C1I 8-PREP" (NACALAT TESQUE, INC.), acetonitrile/water 6/94 followed by concentration by evaporation under reduced pressure and lyophilization, whereby the title compound (156 mg) was obtained as a powder.

Infrared absorption spectrum (KBr) v max cm-': 1756,1687,1633,1592,1454,1386,131312841261,1226,1182.

Nuclear magnetic resonance spectrum (400 M1Hz, D 2 0) 8 PPM: 1 .22(3H,d,J=7.2Hz), I .30(3H,d,J=6.2Hz), 1.57-1 .67(1H~m), 1.70-1 .85(1H,m), 2.12- 2.27(1H,m), 2.25(3H,s), 2.58-2.79(2HKm), 3,03-3.85(1 1H~m), 3.92-4.01(1H~m), 4.18- 4.29(2H,m), (Example 11) (1 R, 5S,6S)-2-[(2S,4S)-2-r(3 S)-3 -Guanidinomethylpyrrolidin- 1-ylcarbonyl]pyrrolidin-4ylthio]-6-[( 1 1 -hydroxyethyll]-I -mnethyl- I -carbapen-2-em-3 -carboxylic acid (Stereoisomer of Exemplified Compound No. 18) OH C3 H H CO H H HCH O N NH 2

CH

3 INH NH

COOH

Doc: 97162p2.doe Doc :7162p.docP79569/FP-9716(PCTI)/tsa-iglEnglish translationl07. 10.98 To a solution of 4-nitrobenzyl 1-hydroxyethyl]- 1methyl-2-(diphenylphosphoryloxy)- 1 -carbapen-2-em-3-carboxylate (874 mg) in anhydrous acetonitrile (9 ml), N,N-diisopropylethylamine (0.256 ml) and a solution of (2S,4S)-4-mercapto- 1 4 -nitrobenzyloxycarbonyl)-2-[(3S)-3-(4-nitrobenzyloxycarbonylguanidinomethyl)pyrrolidin- 1 -ylcarbonyl]pyrrolidine (926 mg) in anhydrous acetonitrile (1.0 ml) were added while stirring in an ice bath. The resulting mixture was stirred overnight at 0 0 C. To the reaction mixture, ethyl acetate was added. The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure.

The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol whereby 4-nitrobenzyl -hydroxyethyl]-1methyl-2-[(2S,4S)- 1 -(4-nitrobenzyloxycarbonyl)-2-[(3S)-3-(4-nitrobenzyloxycarbonylguanidinomethyl)pyrrolidin- 1 -ylcarbonyl]pyrrolidin-4-ylthio]-1-carbapen-2em-3-carboxylate (1.23 g) was obtained.

Infrared absorption spectrum (KBr) v max cm-': 1773,1710,1656,1607,1521,1438,1404,1385,1346,1285,1208,1177,1135,1109,1030.

Nuclear magnetic resonance spectrum (400 MHz, CDCl 3 8 ppm: 1.28(3H,d,J=7.2Hz), 1.34(3H,d,J=6.3Hz), 1.65-1.85(1H,m), 1.90-2.10(2H,m), 2.15- 2.80(5H,m), 3.05-3.70(11Hm), 4.00-4.10(1H,m), 4.10-4.30(2H,m), 4.45-4.60(1Hm), 5.00-5.55(6H,m), 7.40-7.70(6Hm), 8.15-8.30(6H,m).

To a solution of the compound (1.14 which had been obtained in in tetrahydrofuran (38 ml) and water (19 ml), a 10% palladium-carbon catalyst (2.3 g) was added. The resulting mixture was allowed to absorb hydrogen for 1.2 hours while stirring at an external temperature of 30 0 C. The catalyst was then filtered off. The filtrate was washed with ether, followed by concentration by evaporation under reduced pressure. The residue was purified by reversed-phase column chromatography ["Cosmosil 75C18-PREP" (NACALAI TESQUE, INC.), acetonitrile/water 8/92], followed by concentration by evaporation under reduced pressure and lyophilization, whereby the title compound (187 mg) was obtained as a powder.

Nuclear magnetic resonance spectrum (270 MHz, D 2 0, sodium trimethylsilylpropionate-d 4 as internal standard) 5 ppm: 1.25(3H,d,J=7.0Hz), Doc: 97162p2.doc P79569/FP-9716(PCI)/tsa-igEnglish translation07.10.98 1 .33(3H,d,J=6.3Hz), 1.55-1 .90(2HKm), 2. 1O-2.30(1H~m), 2.50-2.90(2H,m), 3.05- 3.35(4H,m), 3.35-3.5O(3HKm), 3.60-3.75(2H,m), 3.78-3 .90(1H,m), 3.90-4. lO(lFIIm), 4.20-4.35(2H,m).

(Example 12) (1 R, 1R)-l1-Hydroxyethyl]-l1-methyl-2-[(2S,4S)-2-[(3R)-3 -(pyrrolidin- 1ylmethyl)pyrrolidin- 1-ylcarbonyllpyrrolidin-4-ylthiol- 1-carbapen-2-em-3 -carboxylic acid (Stereoisomer of Exemplified Compound No. 54) OH

H

3 H H CH3CO CH3

NH

O

COOH

To a solution of 4-nitrobenzyl (1R,5,6S)-2-(diphenylphosphoryloxy)-6- R)-l1-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3-carboxylate (225 mg) in anhydrous dimethylformnamide (3 ml), a solution of (2S,4S)-4-mercapto-1-(4nitrob enzyloxycarbonyl)-2-[(3 R)-3 -(pyrrolidin- 1 -ylmethyl)pyrroli din- 1 ylcarbonyl]pyrroli dine (198 ml) in anhydrous dimethylformamide (1 ml) and diisopropylethylamine (97 [LI) were added in an ice bath. The resulting mixture was stirred for 20 hours at the same temperature. The reaction mixture was concentrated by evaporation under reduced pressure. The residue was dissolved in ethyl acetate. The resulting mixture was washed successively with water and saturated saline solution and dried over anhydrous sodium sulfate. The solvent was then distilled off. The residue was subjected to chromatography through a silica gel column and from the fraction eluted with ethyl acetate/methanol 4-nitrobenzyl (1R,5S,6S)-6-[(1R)-1hydroxyethyl]-l1-methyl-2-[(2S,4S)- 1-(4-nitrobenzyloxycarbonyl)-2-[(3R)-3- (pyrrolidin- 1-ylmethyl)pyrrolidin- 1-ylcarbonyl]pyr-rolidin-4-ylthio]-l1-carbapen-2-em-3 carboxylate (235 mg) was obtained as a powder.

Infrared absorption spectrum (KBr) v max cm-1: 3351, 2970, 1773, 1709, 1697, 1525, 1447, 1246.

Doe: 97162p2.doc Doe: 7162p.docP79569/FP-9716(PCT)/tsa-ig/English translation/07.1O.98 Nuclear magnetic resonance spectrum (270 MHz, CDCI3) 8 ppm: 1.20-1.30(3H, 1.34(3H, d, J=6.1Hz), 1.40-2.10(6H, 2.00-3.00(8H, 3.10-3.80(6H, 3.80- 4.40(4H, 4.50-4.70(1H, 5.00-5.40(3H, 5.40-5.55(1H, 7.40-7.55(2H, m), 7.60-7.70(2H, 8.15-8.25(4H, m).

The compound (161 mg) obtained in was dissolved in tetrahydrofuran (3 ml)-water (1.5 ml), followed by the addition of a 10% palladium-carbon catalyst (320 mg). The resulting mixture was hydrogenated at room temperature for 90 minutes. The catalyst was then filtered off. The filtrate was concentrated by evaporation under reduced pressure to remove tetrahydrofuran. The residue was washed by ether and the water layer was concentrated by evaporation under reduced pressure. The residue was subjected to reversed-phase column chromatography ("Cosmosil 75C18-PREP" produced by NACALAI TESQUE, INC.) and the fraction eluted with acetonitrile/water 2/98 1/9 was lyophilized, whereby the title compound (51 mg) was obtained as a powder.

Infrared absorption spectrum (KBr) v max cm' 1 3378, 1763, 1655, 1593, 1489, 1376.

Nuclear magnetic resonance spectrum (400 MHz, D 2 0) 6 ppm: 1.20(3H, d, J=7.2Hz), 1.28(3H, d, J=6.4Hz), 1.65-1.90(2H, 1.95-2.40(9H, 2.65-2.80(1H, 3.00-3.25(3H, 3.30-3.40(2H, 3.40-3.55(2H, 3.60-4.00(4H, 4.00- 4.10(1H, 4.20-4.30(2H, 4.65-4.75(1H, m).

(Example 13) (1R,5S,6S)-6-[(1R)-l-Hydroxyethyl]-2-[(2S,4S)-2-[(3S)-3-(2hydroxyethylaminomethyl)pyrrolidin-1-lcarbonyllpyrrolidin-4-ylthio]- -methyl-1carbapen-2-em-3-carboxylic acid (Stereoisomer of Exemplified Compound No. 3) OH H H

CH

3 H

CON

-I-H H" N OH

CH

3 NHM Doc: 97162p2.doc P79569/FP-9716(PCTytsa-igEnglish translation/07.10.98 (2 S,4 S)- 2 3

R)-

3 2 -Hydroxyethyl-N-4-nitrobenzyloxycarbonylamino.

methyl)pyrrolidin- 1 -ylcarbonyl]-4-mercapto- 1 4 -nitrob enzyloxycarbonyl)pyrroli dine (214 mg) was treated in a similar manner to that described in Example 12-(1) and to afford the title compound (85.1 mg) as a powder.

infrared absorption spectrum (KBr) v max 3409, 1747, 1644, 1601, 1455, 1386.

Nuclear magnetic resonance spectrum (400 MHz, D 2 0) 5 PPM: 1.22(3K1 d, J=7.2Hz), 1.30(3H1, d, Jk6.4Hz), 1.70-1.95(2K1 in), 2.15-2.35(211 in), 2.60-2.85(2H1, mn), 2.85-2.95(1H in), 3.15-3.30(311 in), 3.30-3.50(511, mn), 3.55-3.90(2H1, in), 3.90- 4.00(1H, in), 4.20-4.40(5H, in).

(Example 14) (1 R, 5S, S,4 S)- 2 3 R)-3-(Carbainoylinethylaininoinethyl)pyrrolidin-. 1 ylcarbonyl]pyrrolidin-4-ylthiol-6-[( IR)-l1-hydroxyethyl] -1-methyll-l1-carbapen-2-em-3carboxylic acid (Stereoisoiner of Exemplified Compound No. 6) OH H H OeH HCH3 H1

CON\~

CH

3 NH k: 0

COOH

S)-3-(N-Carbainoylinethyl-N-4nitrobenzyloxycarbonylaminomethyl)pyrroli din- 1 -ylcarbonyl]-4-inercapto- 1 nitrobenzyloxycarbonyl)pyrrolidine (368 mng) was treated in a similar manner to that described in Example 12-(1) and to give the title compound (15 8 mg) as a powder.

Infrared absorption spectrum (KBr) v max cm-1: 3401, 1754, 1695, 1645, 1597, 1455.

Nuclear magnetic resonance spectrum (400 MlHz, D 2 0) 8 ppm: 1.22(3H1, d, J=7.2Hz), 1.30(3H1, d, J=6.4Hz), 1.70-1.90(111 mn), 1.90-2.05(111, in), 2.20-2.35(111, in), 2.65-2.80(111 in), 2.95-3.10(111 in), 3.10-3.22(211 in), 3.22-3.30(111, in), 3.35- 3.60(4H1, in), 3.60-3.95(5H1, in), 4.00-4.08(111, in), 4.22-4.30(211 in), 4.60-470(11, in).

97162p2.doe 971622.docP79569/FP-9716(PCT)/tsa-ig/English translation/07. 10.98 (Example (IR, 5 S, 6S)- 2 2 S, 4 -(2-Aminoethylaminom ethyl)pyrrol idin- 1ylcarbonyl]pyrrolidin-4-ylthio]-6-[( 1R)-l1-hydroxyethyl] -1-methyl-i -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No. 12) OH3 H COJ H HHH3

NC

CH

3

NH

NN

O

COOH

(2S,4S)-4-Mercapto-l1-( 4 -nitrobenzyloxycarbonyl)-2-[(3 nitrobenzyloxycarbonylamino)ethyl-N4nitrobenzyloxycarbonylaminomethyl)pyrrolidin-1I-ylcarbonyl]pyrroli dine (248 mg) was treated in a similar manner to that described in Example 12-(1) and to afford the title compound (48.5 mg) as a powder.

Infrared absorption spectrum (KBr) v max cm-1: 3370, 1758, 1648, 1603, 1455, 1386.

Nuclear magnetic resonance spectrum (400 MIHz, D 2 0) 5 PPM: 1.22(3H, d, J=7.211z), 1.30(3H1, d, J=6.411z), 1.70-1.90(111 in), 1.90-2.05(111 in), 2.15-2.33(111, in), 2.55-2.70(111 in), 2.97-3.13(4H1, mn), 3.15-3.33(5K1 in), 3.34-3.52(3H, in), 3.57- 3.67(1H, mn), 3.68-3.90(3H, in), 4.2 1-4.30(2H1, in), 4.60-4.67(111 in).

(Example 16) (1 R, 5S,6S)-2-[(2 S, 4 S)--[3R)-3-(2-Diinethylaninoethylaninomethyl)pyrolidin-. 1 ylcarbonylIpyrrolidin-4-ylthiolj-6-r(I1R)-l1-hydroxyethyl]l -methyl-I -carbapen-2-ein-3 carboxylic acid (Stereoisomer of Exemplified Compound No. 14) 97162p2.doe 971622.docP79569/FP-9716(PC'rytsa-ig/English transiationO7. 10.98 S)-3 -(N-2-Dimethylaminoethyl-N-4nitrobenzyloxycarbonylaminomethyl)pyrrolidin- 1 -ylcarbonyl]-4-mercapto- 1 nitrobenzyloxycarbonyl)pyr-rolidine (428 mg) was treated in a similar manner to that described in Example 12-(1) and to give the title compound (109 mg) as a powder.

Nuclear magnetic resonance spectrum (270 MiHz, D 2 0) 8 PPM: 1.22(3H, d, J=7.2Hz), 1.30(3H, d, J=6.3Hz), 1.60-1.90(2K, in), 2.10-2.25(1, in), 2.45-2.60(1H, in), 2.77(31-I 2.79(3H, 2.70-3.00(2H-, in), 3.OO-3.10(1H, in), 3.3 1-3.53(5H, mn), 3.55-3.87(3H, in), 3.88-4.00(lH, in), 4.20-4.30(2H, in), 4.30-4.40(1H, in).

(Example 17) (1 R, 5S,6 S)-6-[II(1R)-l1-Hydroxyethyl]-l1-methyl-2-[(2S,4S)-2-r(3R)-3 -(N-inethyl-N-2methylaminoethylaminoinethyl)pyrrolidin- I -ylcarbonyllpyrrolidin-4-ylthio]- 1 carbapen-2-ein-3-carboxylic acid hydrochloride (hydrochloride of the stereoisoiner of Exemplified Compound No. 3 1) Oe, H 3H H CH 3

CH-

3 I NH N .HC1 O

COOH

2

S,

4

S)-

4 -Mercapto-2-[(3R)-3-[-2-iN-ethyl-N4-nitrobenzyloxycarbonylainino)ethyl-N-inethylaminoinethyl]pyrroli din- 1 -ylcarbonyl]- 1 nitrobenzyloxycarbonyl)pyrrolidine (617 mng) was treated in a similar manner to that described in Example to afford 4-nitrobenzyl hydroxyethyl]-l1-methyl-2-[(2S,4 1-(4-nitrobenzyloxycarbonyl)-2-[(3R)-3-[N-2-(Ninethyl-N- 4 -nitrobenzyloxycarbonylamino)ethyl-N-methylaminonethyl]pyrrolidin- 1ylcarbonyl]pyrrolidin-4-ylthio]-l1-carbapen-2-em-3 -carboxylate (527 mg) as a powder.

The compound (202 mg) obtained in was dissolved in water (1.5 ml), IN hydrochloric acid (0.20 1 ml) and tetrahydrofuran (3 ml), followed by the addition of a palladium-carbon catalyst (400 mg). The resulting mixture was hydrogenated at room temperature for 45 minutes. The catalyst was then filtered off. The filtrate was concentrated by evaporation under reduced pressure to remove tetrahydrofuran. The Doc: 97162p2.doc Doc:9712p2docP79569/FP-9716(PCTy/tsa-ig/English translationl07. 10.98 residue was washed by ether and the water layer was concentrated by evaporation under reduced pressure. The residue was subjected to reversed-phase column chromatography ("Cosmosil 75C 1 8-PREP" produced by NACALAI TESQUE, INC.) and the fraction eluted with water was lyophilized, to afford the title compound (61 mg) as a powder.

Nuclear magnetic resonance spectrum (400 MlHz, D 2 0) 5 PPM: 1.2 1(3H1, d, J=7.211z), 1.29(3H, d, J=6.311z), 1.70-1.87(111 in), 1.96-2.05(IK1 in), 2.22-2.34(1H, in), 2.60-2.69(111 in), 2.75(3K1 2.90(31L, 3.00-3.10(111, in), 3.10-3.35(7H, in), 3.35-3.44(111, in), 3.45-3.60(3H, in), 3.62-3.92(3H, in), 4.01-4.08(1H, in), 4.20- 4.27(2H1, in), 4.65-4.79(1H, in).

(Example 18) (1 R, 5S, S, 4

S)-

2

-I(

3

R)-

3 -(2-Fluoroethylaininomethyl)pyrrolidinylcarbonyl]pyrrolidin-4-ylthio]-6-[( 1R)-l1-hydroxyethyl]- 1-methyl-I -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No. 4) OH C3 H H_ 0N- H H H C 3 H IO

OH

3 K N

NN

0

COOH

S)-3 2 -Fluoroethyl-N-4-nitrobenzyloxycarbonylaminom ethyl)pyrrolidin- 1 -ylcarbonyl]-4-m ercapto- I -(4-nitrob enzyloxycarbonyl)pyrroli dine can be treated in a similar manner to that described in Example 12-(1) and to afford the title compound.

Doe: 97162p2.doe Doc:9712p2docP79S69/FP-9716(PCTytsa-ig/English translationO7.1O.98 (Example 19) (IR, 5 1-Methylguanidinomethyl)pyrrolidin- 1ylcarbonylljpyrrolidin-4-ylthio-6-[( 1R)-l1-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No. 36) OH CH HCH 3 H H N~IO NH 2

CH

3

N

I NH N O

COOH

(2S,4S)-4-Mercapto-2-[(3R)-3 -methyl-2,3 -bis(4nitrob enzyloxycarbonyl)guanidinomethyl]pyrro i din- 1 -ylcarbonyl]- 1 nitrob enzyloxycarbonyl)pyrroli dine (996 mg) was treated in a similar manner to that described in Example 12-(1) and to afford the title compound (149 mg).

Nuclear magnetic resonance spectrum (400 MI-z, D 2 0) 8 PPM: 1.22(3H1, d, J=7.2Hz), 1.30(3H, d, 1=6.4Hz), 1.55-1.70(111 in), 1.70-1.85(111 in), 2.10-2.25(111, in), 2.65-2.80(2K in), 3.09(3K1 3.15-3.30(211, in), 3.35-3.55(6H1, in), 3.70-3.85(211 in), 3.95-4.05(111 in), 4.20-4.30(211 in).

(Example (1 R, S, 6

S)-

2 2

S,

4 S)-2-[(3R)-3-(N-Acetimidoyl..N.methylaminomethyI)pyrrolidin. 1 ylcarbonyllpyrrolidin-4-ylthio-6-r( IR)- 1-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No. OH H Ur CH 3 H HCH

CONH

CH

3

N

NH NH O

COOH

(2S,4S)-4-Mercapto-l1-(4-nitrobenzyloxycarbonyl)-2-[(3R)-3 nitrobenzyloxycarbonylacetimidoyl-N-methylaminomethyl)pyrrolidin 1ylcarbonyl]pyrrolidine can be treated in a similar manner to that described in Example 12-(1) and to afford the title compound.

Doc: 97162p2.doc Dac: 7162p.docP79569IFP-9716(PCT/tsa-ig/English translation/07. 10.98 (Example 21) (1 R, 5 S, S, 4

S)-

2 -r(3R)-3-(N-Formimidoyl.N-methylaminomethyl)pyrrolidin 1ylcarbonyl]pyrrolidin-4-ylthio]-6- 1 -hydroxyethyl]- I1-methyl- I -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No. 34) OH H 3 H H CH 3 H HHH :CON D

CH

3 S ,N I NH N o

COOH

(2 S,4 S)-4-Mercapto-lI-(4-nitrobenzyloxycarbonyl)-2-[(3R)-3 nitrobenzyloxycarbonylformimidoyl-N-methylaminomethyl)pyrrolidinylcarbonyl]pyrrolidine can be treated in a similar manner to that described in Example 12-(1) and to give the title compound.

(Example 22) iR)-l1-Hydroxyethyl]-l1-methyl-2-[(2S,4S)-2-r(3R)-3-(2,2,2trifluoroethylaminomethyl)pyrrolidin-.1 -ylcarbonyl]pyrrolidin-4-ylthio]-l1-carbapen-2em-3-carboxylic acid (Stereoisomer of Exemplified Compound No. OH C3 H ON H He, H H CH 3 I- ICON

S

CH

3

N

O

COOH

(2S,4S)-4-Mercapto-l1-( 4 -nitrobenzyloxycarbonyl)-2-[(3 nitrobenzyloxycarbony1-N-2,2,2-trifluoroethylaminomethyl]pyrroli din- 1ylcarbonyl]pyrrolidine can be treated in a similar manner to that described in Example 12-(1) and to give the title compound.

Vr A/7' O ~Dm: 97162p2.doe P79569/FP-9716(PCT)tsa-ig/English translation/07. 10.98 63 (Example 23) (I R, 5S, -Form imidoylaminomethylpyrrol idin- 1yl carbonyllpyrrolidin-4-ylthio]-6-[( I R 1 -hydroxyethyl]-I 1-m ethyl- I -carbapen-2-em- 3-carboxylic acid (Stereoisomer of Exemplified Compound No. 16) OH H H

OH

3 H ICON()"" H

OH

3

NH

N

NHH

(2 S, 4 S) -4-Mercapto 1 -ni1trob en zyl ox ycarb on y1) (3 3 nitrob enzyl oxycarbonyl form imi doyl am inom ethylI)pyrro 1idin- I -yl carbonyl] pyrrol id ine can be treated in a similar manner to that described in Example 12-(1) and to afford the title compound.

(Example 24) 5S, 6

S)-

2 -[(2S,4S)-2-[(3R)-3-Guanidinomethylpyrrolidin-1 -ylcarbonyl]- 1m ethyl pyrro Ii din-4-ylthio] 1 1 -hydroxyethyl]- 1 -m ethyl- I -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No. 79) OH H H H He, HH H3 %CN ,N NH 2

-OH

3 N

NH

OH

3 0

GOH

2

S,

4

S)-

2 -[(3R)-3-[2,3-bis(4-nitrobenzyloxycarbonyl) guani dinomnethyl] pyrrolidin-1I-ylcarbonyl]-4-mercapto- I -m ethylpyn-olidine can be treated in a similar manner to that described in Example and to give title compound.

(Example

S,

6

S)-

2 2 S,4S)-2-[(3R)-3-Acetimidoylaminomethylpyrrolidin 1 -ylcarbonyl]- 1methylpyrrolidin-4-ylthio]-6-[( 1R)-l1-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No. 78) OH HH H, HHCH 3 H ICON\H 3

CH

3 N\ 3

NH

O

COOH

(2 S,4 S)-4-Mercapto-l1-methyl-2-[(3R)-3-(N-4-nitrobenzyloxycarbonyl.

acetimidoyl-N-methylaminomethyl)pyrrolidin-1 -ylcarbonyl] pyrrolidine can be treated in a similar manner to that described in Example and to afford the title compound.

(Example 26) (1R 5 S,1 6

S)-

2 2

S,

4

S)-

2 -[(3R)-3-Formimidoylaminomethylpyrrolidin-.1 -ylcarbonyl]- 1methylpyrrolidin-4-ylthio 1 1 -hydroxyethyl 1- 1 -m ethyl- I -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No. 77) OH C3 H H H H H NH IO

S

N I\ H3NH 0

COOH

(2S,4S)-4-mercapto- I -methyl- 2 -[(3R)-3-(4-nitrobenzyloxycarbonyl.

formimidoylaminomethyl)pyrrolidin- 1-ylcarbonyl]pyrrolidine can be treated in a similar manner to that described in Example and to give the title compound.

Doe: 97162p2.doc Doc: 7162p.docP79569/FP-9716(PCT)tsa-ig/English translation/07.10.98 (Example 27) (1 R, 5S, 6S)-2-[(2S,4 1-Methylguanidinomethyl)pyrrol idin- 1-ylcarbonyl]- 1 -methylpyrrolidin-4-ylthio]-6-[( 1R)-l1-hydroxyethyl]- 1-methyl-i -carbapen-2-em-3 carboxylic acid (Stereoisomer of Exemplified Compound No, 96) OH H 0CH 3 He,, H H 'I N

ION

CH

3 S y H CH 3

N

o

COOH

3-Bis(4-nitrobenzyloxycarbonyl)- 1methylguanidinomethyl]pyrrolidin- 1-ylcarbonyl]-4-mercapto-l1-methylpyrrolidine can be treated in a similar manner to that described in Example and to afford the title.

(Example 28) (I R, 5 S, S, 4

S)-

2 3

R)-

3 -(N-Acetimidoyl-N-methylaminomethyl)pyrroli din-.. 1ylcarbonyl]-l1-methylpyrrolidin-4-ylthio]-6- II(1R)-l1-hydroxyethyl]- 1-methyl-i carbapen-2-em-3 -carboxylic acid (Stereoisomer of Exemplified Compound No. OH H

CH

3 H H SH C NC O

CH

3 CH3 K: N\CH O COOH

H

(2S,4S)-4-Mercapto- 1 -methyl-2-[(3R)-3-(N-4nitrobenzyloxycarbonylacetimidoyl-N-methylaminomethyl)pyruolidin-l-1 ylcarbonyl]pyrrolidine can be treated in a similar manner to that described in Example and to give the title compound.

Doc: 97162p2.doc Dac:9162p.docP79569/FP-9716(PCTytsa-igfinglish translationO7. 10.98 (Example 29) (I R, 5 S, S, 4S)-2- -(N-Form im idoyl-N-methylaminom ethyl)pyrroli din-. 1ylcarbonyl]- 1 -methylpyrrolidin-4-ylthio]-6- 1 -hydroxyethyl]- I1-methyl- I carbapen-2-em-3-carboxylic acid (Stereoisomer of Exemplified Compound No. 94) OH H 3 H H CH 3 HI: HC N

CH

3 N-N H NH o COOH

CH

(2S,4S)-4-Mercapto- 1 -methyl-2-[(3R)-3-(N-4nitrobenzyloxycarbonylformnimidoyl-N-methylaminomethyl)pyrrolidinylcarbonyl]pyrrolidine can be treated in a similar manner to that described in Example and to afford the title compound.

(Example (1 R, 5S,6 1R)-l1-Hydroxyethyl]-l1-methyl-2-[(2S,4 m ethyl aminom ethylpyrrolidin- 1 -yl carbonyl]pyrrolidin-4-ylthio]- 1 -carb apen-2-em-3 carboxylic acid hydrochloride (Hydrochloride of the stereoisomer of Exemplified Compound No. 1) OH H H HCH HOND ~,,,,NHCH 3

HCI

CH

3

-N

O

COOH

To an aqueous solution (2 ml) of (1R, 5 S, IR)-l1-hydroxyethyl]- 1-methyl- 2-[(2S,4 S)-2-[(3R)-3-methylaminomethylpyrrolidin-.1 -ylcarbonyl]pyrrolidin-4-ylthio]- I -carbapen-2-em-3-carboxylic acid (59.7 mg), obtained in Example 3, IN hydrochloric acid 122 ml) was added. The resulting aqueous solution was subjected to reversedphase column chromatography ("Cosmosil 75C1I8-PREP" produced by NACALAI TESQUE, INC.) and from the fraction eluted with water, the title compound (46 mg) was obtained as a powder.

Doe: 97162p2.doc Doc:9162p.docP79569IFP-9716(PCTy/tsa-ig/English translationl07. 10.98 Infrared absorption spectrum (KBr) v max 3409, 1757, 1634, 1598, 1456, 1386.

Nuclear magnetic resonance spectrum (400 MHz, D 2 0) 8 ppm: 1.21(3H, d, J=7.2Hz), 1.29(3H, d, J=6.3Hz), 1.70-1.87(1H, 1.96-2.05(1H, 2.20-2.32(1H, 2.60-2.70(1H, 2.75(3H, 3.00-3.10(1H, 3.10-3.35(3H, 3.35-3.45(1H 3.45-3.50(3H 3.60-3.90(3H, 4.02-4.09(1H, 4.21-4.28(2H, 4.66- 4.78(1H, m).

(Referential Example 1) (2S,4S)-4-Mercapto-l-methyl-2-[(3R)-3-(4-nitrobenzyloxycarbonylaminomethyl)pyrrolidin-1 -ylcarbonyl]pyrrolidine To a solution (240 ml) of (3S)-3-hydroxymethyl-l-[(1R)-lphenylethyl]pyrrolidine (11.5 g) in ethanol, a palladium hydroxide-carbon catalyst (11.6 g) was added. The resulting mixture was allowed to absorb hydrogen for 3 hours while stirring at an external temperature of 40 0 C. The catalyst was then filtered off, followed by concentration by evaporation under reduced pressure. Into the residue (6.01 g), acetonitrile (60 ml) was poured and then di-tert-butyl carbonate (14 ml) was added to the resulting mixture in an ice bath. After the temperature of the reaction mixture was allowed to rise to room temperature, the mixture was stirred for one hour. Saturated saline solution was then poured into the reaction mixture to terminate the reaction, followed by extraction three times with ethyl acetate. The combined organic layer was washed with saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (ethyl acetate), whereby (3S)-1-tertbutoxycarbonyl-3-hydroxymethylpyrrolidine (8.66 g) was obtained.

Optical rotation: [a]D 25 =-16.50 CHC13).

Infrared absorption spectrum (Liquid film) v max cm" 1 3432,1698,1675,1479,1454,1418,1367.

Doc: 97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English translation/07.1O.98 Nuclear magnetic resonance spectrum (270 MHz, CDC13) 6 ppm: 1.46(9H,s), 1.61-1.80(1H,m), 1.92-2.30(2H,m), 2.37-2.47(1H,m), 3.07-3.17(1H,m), 3.30- 3.69(5H,m).

To a solution of the compound (1.30 g) obtained in Referential Example 1in tetrahydrofuran (13 ml), triethylamine (0.99 ml) and methanesulfonyl chloride (0.55 ml) were successively added in an ice bath, followed by stirring for one hour. Into the reaction mixture, saturated saline solution was poured to terminate the reaction, followed by extraction three times with ethyl acetate. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure. Into the residue (1.83 N,Ndimethylformamide (20 ml) was poured, to which sodium azide (1.26 g) was added.

The resulting mixture was stirred at 80 0 C for 1.5 hours. Into the reaction mixture, saturated saline solution was poured to terminate the reaction, followed by extraction three times with diethyl ether. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (n-hexane/ethyl acetate whereby (3S)-3azidomethyl-1-tert-butoxycarbonylpyrrolidine (1.39 g) was obtained.

To a solution of the compound (1.23 g) obtained in Referential Example 1in acetonitrile (13 ml), triphenylphosphine (1.50 g) was added and the resulting mixture was refluxed for one hour. To the reaction mixture, 4-nitrobenzyl chloroformate (1.52 g) and a IN aqueous sodium hydroxide solution (7 ml) were successively added. After the temperature was allowed to rise to room temperature, the mixture was stirred for 30 minutes. The reaction mixture was diluted with water and then extracted three times with methylene chloride. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (n-hexane/ethyl acetate whereby (3R)-1-tert-butoxycarbonyl-3-(4-nitrobenzyloxycarbonylaminomethyl)pyrrolidine (1.97 g) was obtained.

Optical rotation: [a]D 2 5 =-14.30 CHCl 3 Doc: 97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English translation/07.1O.98 Infrared absorption spectrum (Liquid film) v max cm-: 3326,1727,1683,1524,1413,1348,1250.

Nuclear magnetic resonance spectrum (270 MHz, CDC13) 6 ppm: 1.45(9H,s), 1.55-1.66(1H,m), 1.92-2.03(1H,m), 2.35-2.45(1H,m), 2.95-3.55(6H,m), 5.18(1H,br.s), 5.20(2H,s), 7.51(2H,d,J=8.6Hz), 8.22(2H,d,J=8.6Hz).

To a solution (20 ml) of the compound (1.90 g) obtained in Referential Example in methylene chloride, trifluoroacetic acid (3.9 ml) was added in an ice bath. After the temperature of the reaction mixture was allowed to rise to room temperature, the mixture was stirred for 2 hours. The reaction mixture was diluted with methylene chloride, followed by extraction three times with water. Into the combined water layers, a IN aqueous sodium hydroxide solution (60 ml) was poured to make the solution alkaline. The mixture was extracted three times with methylene chloride. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure, whereby 4 -nitrobenzyloxycarbonylaminomethyl)pyrrolidine (1.38 g) was obtained as the crude product.

To a solution of (2S,4S)-4-(4-methoxybenzylthio)-l-methyl-2pyrrolidinecarboxylic acid (0.69 g) in tetrahydrofuran (7 ml), N,Ndiisopropylethylamine (0.43 ml) and pivaloyl chloride (0.30 ml) were added in an ice bath. The resulting mixture was stirred at 0°C for 10 minutes. To the reaction mixture, a solution of a mixture of the compound (0.68 g) obtained in Referential Example 1-(4) and N,N-diisopropylethylamine (0.43 ml) in acetonitrile (8 ml) was added and the resulting mixture was stirred at 0°C for 30 minutes. Into the reaction mixture, saturated saline solution was poured to terminate the reaction, followed by extraction three times with ethyl acetate. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol whereby (2S,4S)-4-(4-methoxybenzylthio)-lmethyl- 2 -[(3R)-3-(4-nitrobenzyloxycarbonylaminomethyl)pyrrolidin-1ylcarbonyl]pyrrolidine (0.73 g) was obtained.

Doc: 97162p2.doc P79569/FP-9716(PCI)/tsa-ig/English translation/07. 10.98 Infrared absorption spectrum (Liquid film) v max cm 1722,1639,1610,1585,1513,1445,1373,1347,1322,1302,1247,1177,1144,1109.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 8 ppm: 1.50- 2.62(5H,m), 2.29,2.31(3H,sx2), 3.02-3.83(10H,m), 3.69,3.70(2H,sx2), 3.79(3H,s), 4.98- 5.18(1H,m), 5.19(2H,s), 6.83(2H,d,J=6.6Hz), 7.21(2H,d,J=6.6Hz), 7.50(2H,d,J=8.6Hz), 8.22(2H,d,J=8.6Hz).

To a solution of the compound (0.65 g) obtained in Referential Example 1in a mixture of anisole (0.65 ml) and trifluoroacetic acid (6.5 ml), trifluoromethanesulfonic acid (0.27 ml) was added while stirring in an ice bath. After the temperature of the reaction mixture was allowed to rise to room temperature, the mixture was stirred for one hour. Trifluoroacetic acid was distilled off under reduced pressure and the residue was washed with n-hexane and diethyl ether to remove anisole.

Ethyl acetate was added to the residue. The resulting mixture was washed with a saturated aqueous solution of sodium bicarbonate and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure, whereby the title compound (0.54 g) was obtained.

[Another method for synthesis of the title compound] In anhydrous pyridine (100 ml), (3R)-1-tert-butoxycarbonyl-3-pyrrolidinole (10.0 g) was dissolved. To the resulting solution, dimethylaminopyridine (652 mg) and p-toluenesulfonyl chloride (15.3 g) was added in an ice bath, followed by stirring for 48 hours in an ice bath. The solvent was removed under reduced pressure. The residue was dissolved in methylene chloride (200 ml), followed by washing with water. After the water layer was extracted twice with methylene chloride, all the organic layers were washed with saturated saline solution. The organic layers were dried over anhydrous magnesium sulfate and subjected to concentration by evaporation under reduced pressure, whereby 24.2 g of the crude product were obtained. The resulting crude product was purified by chromatography through a silica gel column (eluent: methylene chloride/acetonitrile 40/1), whereby (3R)-l-tert-butoxycarbonyl-3-ptoluenesulfonyloxypyrrolidine (16.8 g) was obtained.

Doc: 97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English translation/07.1 O.98 Nuclear magnetic resonance spectrum (270 MHz, CDC1 3 8 ppm: 1.43(9H,s), 2.25-1.90(2H,m), 2.46(3H,s), 3.60-3.30(4H,m), 5.05(1H,m), 7.35(2H,d,J=7.9Hz), 7.79(2H,d,J=7.9Hz).

The compound (12.2 g) obtained in was dissolved in anhydrous acetonitrile (122 ml). To the solution, 1,8-diazabicyclo[5,4,0]-7-undecene (8.02 ml) and acetone cyanohydrin (6.53 ml) were added, followed by heating under reflux for hours. The reaction mixture was diluted with ethyl acetate (1 liter) and washed five times with water (200 ml) and once with a saturated aqueous solution of ammonium chloride (200 ml). The organic layer was dried over anhydrous magnesium sulfate and concentrated by evaporation under reduced pressure, whereby the crude product (9.10 g) was obtained. The resulting crude product was purified by column chromatography (eluent: benzene/ethyl acetate 17/1), whereby (3R)-1-tert-butoxycarbonyl-3cyanopyrrolidine (4.41 g) was obtained.

Nuclear magnetic resonance spectrum (270 MHz, CDC1 3 5 ppm: 1.47(9H,s),2.35-2.15(2H,m),3.15-3.00(1H,m),3.75-3.35(4H,m).

In anhydrous tetrahydrofuran (21 ml), the compound (2.10 g) obtained in (8) was dissolved. To the solution, lithium aluminum hydride (2.03 g) was added in an ice bath, followed by stirring in an ice bath for 20 minutes and at room temperature for minutes. To the reaction mixture, tetrahydrofuran (40 ml) and water (4.6 ml) were added and the resulting mixture was stirred at room temperature for 10 minutes. The solvent was removed under reduced pressure. To the residue, methylene chloride (250 ml) and anhydrous sodium sulfate (24 g) were added and the resulting mixture was stirred at room temperature for one hour. After filtration, the solvent was distilled off under reduced pressure. The residue was purified by chromatography through a silica gel column (eluent: ethyl acetate/methanol whereby (3R)-3-aminomethyl-l-tertbutoxycarbonylpyrrolidine (703 mg) was obtained.

Nuclear magnetic resonance spectrum (270 MHz, CDC13) 8 ppm: 1.30- 1.50(2H,br.s), 1.46(9H,s), 1.50-1.65(1H,m), 1.90-2.10(1H,m), 2.15-2.30(1H,m), 2.65- 2.80(2H,m), 2.90-3.10(1H,m), 3.20-3.60(3H,m).

In anhydrous acetonitrile (5.5 ml), the compound (550 mg) obtained in (9) was dissolved. To the solution, diisopropylethylamine (0.575 ml) and p-nitrobenzyl Doe: 97162p2.doc P79569/FP-9716(PCTytsa-igEnglish translation/07.1O.98 chloroformate (711 mg) were added in an ice bath, followed by stirring for 5 minutes in an ice bath. The solvent was removed under reduced pressure. The residue was dissolved in methylene chloride (50 ml) and the resulting solution was washed with water. The water layer was extracted twice with methylene chloride. All the organic layers were washed with saturated saline solution. The organic layers were dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure. The residue was then purified by chromatography through a silica gel column (eluent: benzene/acetonitrile whereby (3R)-1-tert-butoxycarbonyl-3-(4nitrobenzyloxycarbonylaminomethyl)pyrrolidine (599 mg) was obtained. The resulting product coincided completely with that obtained in in data of optical rotation, infrared absorption spectrum and nuclear magnetic resonance spectrum.

(Referential Example 2) (2S,4S)-4-Mercapto-1-methyl-2-[(3S)-3-(4-nitrobenzyloxycarbonylaminomethyl)pyrrolidin-1-ylcarbonyl]pyrrolidine To a solution (250 ml) of (3R)-3-hydroxymethyl-l-[(1R)-lphenylethyl]pyrrolidine (12.6 g) in ethanol, a palladium hydroxide-carbon catalyst (12.5 g) was added. The resulting mixture was allowed to absorb hydrogen for 5 hours while stirring at an external temperature of 40 0 C. The catalyst was filtered off, followed by concentration by evaporation under reduced pressure. Into the residue (7.08 g), acetonitrile (70 ml) was poured, to which di-tert-butylcarbonate (16 ml) was added in an ice bath. After the temperature of the reaction mixture was allowed to rise back to room temperature, the reaction mixture was stirred for one hour. The reaction mixture was treated in a similar manner to that described in Referential Example whereby (3R)-l-tert-butoxycarbonyl-3-hydroxymethylpyrrolidine (10.6 g) was obtained.

Optical rotation: [o]D 25 +16.70 CHCl 3 Infrared absorption spectrum (Liquid film) v max cm" 1 3434,1698,1675,1479,1454,1418,1367.

Doc: 97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English translation/07.10.98 Nuclear magnetic resonance spectrum (270 MHz, CDC13) 5 ppm: 1.

4 6 9 H,s), 1.61-1.80(2H,m), 1.92-2.30(1H,m), 2.37-2.47(1H,m), 3.07-3.17(1H,m), 3.30- 3.69(5H,m).

To a solution of the compound (1.55 g) obtained in Referential Example 2in tetrahydrofuran (16 ml), triethylamine (1.18 ml) and methanesulfonyl chloride (0.66 ml) were successively added in an ice bath, followed by stirring for one hour. Into the reaction mixture, saturated saline solution was poured to terminate the reaction. The resulting mixture was extracted three times with ethyl acetate. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure. Into the residue (2.19 g), N,N-dimethylformamide (20 ml) was poured, followed by the addition of sodium azide (1.50 The mixture was stirred at 80 0 C for 1.5 hours. The reaction mixture was treated in a similar manner to that described in Referential Example whereby (3R)-3-azidomethyl- 1-tert-butoxycarbonylpyrrolidine (1.43 g) was obtained.

To a solution of the compound (1.00 g) obtained in Referential Example 2in acetonitrile (10 ml), triphenylphosphine (1.22 g) was added and the resulting mixture was refluxed for one hour. To the reaction mixture, 4-nitrobenzyl chloroformate (1.24 g) and a 1N aqueous sodium hydroxide solution (6 ml) were successively added in an ice bath. After the temperature was allowed to rise to room temperature, the mixture was stirred for 30 minutes. The reaction mixture was treated in a similar manner to that described in Referential Example whereby (3S)-l-tertbutoxycarbonyl-3-(4-nitrobenzyloxycarbonylaminomethyl)pyrrolidine (1.51 g) was obtained.

Optical rotation: [a]D 2 5 +14.50 CHCl 3 Infrared absorption spectrum (Liquid film) v max cm" 1 3325,1726,1682,1524,1414,1348,1249.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 6 ppm: 1.45(9H,s), 1.55-1.66(1H,m), 1.92-2.03(1H,m), 2.35-2.45(1H,m), 2.95-3.55(6H,m), 5.06(1H,br.s), 5.19(2H,s), 7.51(2H,d,J=8.6Hz), 8.22(2H,d,J=8.6Hz).

To a solution of the compound (1.51 g) obtained in Referential Example 2in methylene chloride (17 ml), trifluoroacetic acid (3.1 ml) was added in an ice bath.

Doc: 97162p2.doc P79569/FP-9716(P T)tsa-ig/Engish translation/07.10.98 After the temperature of the reaction mixture was allowed to rise to room temperature, the mixture was stirred for two hours. The reaction mixture was treated in a similar manner to that described in Referential Example nitrobenzyloxycarbonylaminomethyl)pyrrolidine (1.16 g) was obtained as a crude product.

Nuclear magnetic resonance spectrum (270 MHz, CDCI 3 5 ppm: 1.42- 1.60(1H,m), 1.90-2.07(1H,m), 2.27-2.50(1H,m), 2.52-3.28(7H,m), 5.13-5.31(1H,m), 5.19(2H,s), 7.51(2H,d,J=8.6Hz), 8.22(2H,d,J=8.6Hz).

To a solution of (2S,4S)-4-(4-methoxybenzylthio)-l-methyl-2pyrrolidinecarboxylic acid (0.70 g) in tetrahydrofuran (7 ml), N,Ndiisopropylethylamine (0.43 ml) and pivaloyl chloride (0.31 ml) were added in an ice bath. The resulting mixture was stirred at 0°C for 10 minutes. To the reaction mixture, a solution of a mixture of the compound (0.70 g) obtained in Referential Example 2-(4) and N,N-diisopropylethylamine (0.43 ml) in acetonitrile (8 ml) was added, followed by stirring at 0°C for 30 minutes. The reaction mixture was treated in a similar manner to that described in Referential Example whereby (2S,4S)-4-(4-methoxybenzylthio)- 1-methyl-2-[(3S)-3-(4-nitrobenzyloxycarbonylaminomethyl)pyrrolidin-1ylcarbonyl]pyrrolidine (0.75 g) was obtained.

Infrared absorption spectrum (Liquid film) v max cm 1 1722,1639,1610,1585,1513,1445,1347,1302,1248,1176,1144,1120,1109.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 5 ppm: 1.52- 2.61(5H,m), 2.30(3H,s), 3.03-3.78(10H,m), 3.70(2H,s), 3.79,3.80(3H,sx2), 5.04- 5.05(1H,m), 5.19(2H,s), 6.83,6.84(2H,dx2,J=8.6Hz), 7.20,7.22(2H,d 2,J=8.6Hz), 7.51(2H,dxJ=8.6Hz), 8.22(2H,d,J=8.6Hz).

To a solution of the compound (0.67 g) obtained in Referential Example 2in a mixture of anisole (0.67 ml) and trifluoroacetic acid (6.7 ml), trifluoromethanesulfonic acid (0.27 ml) was added while stirring in an ice bath. The temperature of the reaction mixture was allowed to rise to room temperature, followed by stirring for one hour. The reaction mixture was treated in a similar manner to that described in Referential Example whereby the title compound (0.57 g) was obtained.

Doc: 97162p2.doc P79569/FP-9716(PCI)tsa-igEnglisb translation/0 7 .1O.9 8 (Referential Example 3) (2S,4S)-4-Mercapto-2-[(3 S)-3-(N-methyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl)pyrrolidin- 1 -ylcarbonyl]- 1-(4-nitrobenzyloxycarbonyl)pyrrolidine To a solution of the compound (0.82 g) obtained in Referential Example 1in tetrahydrofuran (8 ml), triethylamine (0.62 ml) and methanesulfonyl chloride (0.35 ml) were successively added. The resulting mixture was stirred for one hour. Into the reaction mixture, saturated saline solution was poured to terminate the reaction, followed by extraction three times with ethyl acetate. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure.

To the residue (1.16 a 40% methylamine-methanol solution (18 ml) was added and the mixture was heated in a pressure bottle at 100 0 C for 4 hours. After the temperature of the reaction mixture was allowed to lower to room temperature, the mixture was concentrated by evaporation under reduced pressure. To a solution of the residue (1.32 g) in acetonitrile (15 ml), N,N-diisopropylethylamine (1.4 ml) and 4nitrobenzyl chloroformate (1.73 g) were added in an ice bath, followed by stirring at 0°C for 2 hours. Saturated saline solution was poured into the reaction mixture to terminate the reaction and the resulting mixture was extracted three times with ethyl acetate. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure.

The residue was purified by chromatography through a silica gel column (nhexane/ethyl acetate 4/6 whereby (3R)-1-tert-butoxycarbonyl-3-[N-methyl-N- (4-nitrobenzyloxycarbonyl)aminomethyl]pyrrolidine (1.42 g) was obtained.

Optical rotation: [a]D 2 5 -6.90 CHC13).

Infrared absorption spectrum (Liquid film) v max cm 1 1696,1608,1524,1480,1455,1404,1366,1347,1293,1255,1211,1191,1170,1152,1125.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 8 ppm: 1.45(9H,s), 1.56-1.68(1H,m), 1.89-2.01(1H,m), 2.45-2.55(1H,m), 2.98(3H,s), 2.98-3.10(1H,m), 3.27-3.57(5H,m), 5.23(2H,s), 7.51(2H,d,J=8.6Hz), 8.23(2H,d,J=8.6Hz).

Doc: 97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English translation/07. 10.98 To a solution of the compound (1.96 g) obtained in Referential Example 3in methylene chloride (25 ml), trifluoroacetic acid (3.8 ml) was added in an ice bath, followed by stirring at room temperature for 2 hours. The reaction mixture was treated in a similar manner to that described in Referential Example whereby methyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl]pyrrolidine (1.55 g) was obtained.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 6 ppm: 1.30- 1.51(1H,m), 1.81-1.97(1H,m), 2.20-2.69(3H,m), 2.85-3.06(6H,m), 3.20-3.35(2H,m), 5.22(2H,s), 7.51(2H,d,J=8.6Hz), 8.22(2H,d,J=8.6Hz).

To a solution of (2S,4S)-4-(4-methoxybenzylthio)-1-(4nitrobenzyloxycarbonyl)-2-pyrrolidinecarboxylic acid (1.52 g) in tetrahydrofuran ml), N,N-diisopropylethylamine (0.59 ml) and pivaloyl chloride (0.42 ml) were added in an ice bath. The resulting mixture was stirred at 0 C for 10 minutes. To the reaction mixture, a solution of a mixture of the compound (1.00 g) obtained in Referential Example and N,N-diisopropylethylamine (0.59 ml) in acetonitrile (15 ml) was added and the resulting mixture was stirred at 0 C for 30 minutes. The reaction mixture was treated in a similar manner to that described in Referential Example whereby (2S,4S)-4-(4-methoxybenzylthio)-2-[(3S)-3-(N-methyl-N-(4nitrobenzyloxycarbonyl)aminomethyl)pyrrolidin-1 -ylcarbonyl]-1 nitrobenzyloxycarbonyl)pyrrolidine (2.08 g) was obtained.

Infrared absorption spectrum (KBr) v max cm" 1706,1654,1608,1585,1520,1439,1403,1346,1299,1249,1210,1194,1175,1149,1110.

Nuclear magnetic resonance spectrum (270 MHz, CDC1 3 8 ppm: 1.50- 2.70(5H,m), 2.92-4.08(12H,m), 3.74(2H,s), 3.80,3.81(3H,sx2), 4.22-4.45(1H,m), 4.98- 5.33(4H,m), 6.87(2H,d,J=8.6Hz), 7.22-7.56(6H,m), 8.20-8.30(4H,m).

To a solution of the compound (2.03 g) obtained in Referential Example 3in a mixture of anisole (2.0 ml) and trifluoroacetic acid (20 ml), trifluoromethanesulfonic acid (0.62 ml) was added while stirring in an ice bath. The resulting mixture was then stirred at room temperature for one hour. The reaction mixture was treated in a similar manner to that described in Referential Example whereby the title compound (1.92 g) was obtained.

97162p2.doc P79569/FP-9716(PCI)/tsaigEnglish translation/0l. 10.98 [Another method for a synthesis of the title compound] To a solution of (2S,4R)-4-hydroxy-1-(4-nitrobenzyloxycarbonyl)proline (4.65 g) in anhydrous dimethylformamide (60 ml), (3S)-3-[N-methyl-N-(4nitrobenzyloxycarbonyl)aminomethyl]pyrrolidine hydrochloride (4.95 g), diisopropylethylamine (5.23 ml), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (4.03 g) and 1-hydroxybenzotriazole (2.23 g) were added. The resulting mixture was stirred at room temperature for 10 hours. The reaction mixture was concentrated by evaporation under reduced pressure. The residue was dissolved in ethyl acetate. The resulting solution was washed with water and the organic layer was dried over anhydrous sodium sulfate. The solvent was distilled off and the residue was subjected to chromatography through a silica gel column. From the fractions eluted with ethyl acetate/methanol 9/1, (2S,4R)-4-hydroxy-2-[(3S)-3-[N-methyl-N-(4nitrobenzyloxycarbonyl)aminomethyl]pyrrolidin-1-ylcarbonyl]- 1 nitrobenzyloxycarbonyl)pyrrolidine (8.70 g) was obtained as a powder.

Infrared absorption spectrum (KBr) v max cm' 1 3402, 1706, 1654, 1607, 1522, 1436, 1346.

Nuclear magnetic resonance spectrum (400 MHz, DMSO-d 6 8 ppm: 1.30- 2.20(3H, 2.30-2.60(1H, 2.80-3.00(3H, 3.00-3.80(10H 4.20-4.40(1H, 4.40-4.60(1H, 5.00-5.30(4H, 7.50-7.70(4H, 8.10-8.30(4H, m).

The compound (8.70 g) obtained in was dissolved in anhydrous acetonitrile (87 ml). To the solution, triethylamine (2.72 ml) and methanesulfonyl chloride (1.34 ml) were added in an ice bath, followed by stirring at the same temperature for 5 minutes. The reaction mixture was concentrated by evaporation under reduced pressure. The residue was dissolved in ethyl acetate. The resulting solution was washed successively with water and saturated saline solution and then dried over anhydrous sodium sulfate. The residue was subjected to chromatography through a silica gel column. From the fractions eluted with ethyl acetate/methanol 18/1 14/1, (2S,4R)-4-methanesulfonyloxy-2-[(3S)-3-[N-methyl-N-(4nitrobenzyloxycarbonyl)aminomethyl] pyrrolidin-1-ylcarbonyl]-l-(4nitrobenzyloxycarbonyl)pyrrolidine (9.33 g) was obtained.

97162p2.doc P79569/FP-9716(PCT)/tsa-igfEnglish translation/07. 10.98 Infrared absorption spectrum (KBr) v max cm' 1 1706, 1652, 1607, 1522, 1441, 1405, 1347.

Nuclear magnetic resonance spectrum (400 MHz, DMSO-d 6 6 ppm: 1.50- 2.20(3H, 2.30-2.80(3H, 2.90-3.05(3H, 3.08(3H, 3.10-3.70(6H, 3.80- 4.10(2H, 4.50-4.70(1H, 5.20-5.40(4H, 7.40-7.55(4H, 8.15-8.25(4H, m).

The compound (2.00 g) obtained in was dissolved in anhydrous dimethylformamide (25 ml). To the solution, potassium thioacetate (520 mg) was added and the resulting mixture was stirred at 75 0 C for 1.5 hours. After cooling to room temperature, the reaction mixture was diluted with ethyl acetate, washed with water and saturated saline solution and then dried over anhydrous sodium sulfate. The residue was subjected to chromatography through a silica gel column. From the fractions eluted with ethyl acetate/methanol 100/1, (2S,4S)-4-acetylthio-2-[(3S)-3-[Nmethyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl]pyrrolidin- -ylcarbonyl]- 1 nitrobenzyloxycarbonyl)pyrrolidine (1.60 g) was obtained.

Infrared absorption spectrum (KBr) v max cm'1: 1705, 1654, 1607, 1522, 1437, 1404, 1346.

Nuclear magnetic resonance spectrum (400 MHz, DMSO-d 6 8 ppm: 1.70- 2.10(3H, 2.34(3H, 2.40-2.70(2H, 2.90-3.10(3H, 3.10-3.70(6H, 3.80- 4.10(2H, 4.11(1H, t, J=9.1Hz), 4.40-4.60(1H, 5.00-5.40(4H, 7.40-7.60(4H, 8.15-8.30(4H, m).

The compound (1.48 g) obtained in was dissolved in methanol (30 ml) and methylene chloride (4.4 ml). To the resulting solution, a 1N sodium methoxide solution in methanol (2.3 ml) was added in an ice bath, followed by stirring at the same temperature for 10 minutes. To the reaction mixture, IN hydrochloric acid (2.4 ml) was added and the resulting mixture was concentrated by evaporation under reduced pressure. The residue was dissolved in ethyl acetate. The resulting solution was washed successively with water and saturated saline solution and dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure, whereby the title compound (1.34 g) was obtained as a powder. The resulting compound coincided with that obtained in in infrared absorption spectrum and nuclear magnetic resonance spectrum.

97162p2.doc P795691FP-9716(PCT)tsa-ig/English translation/07.10.98 (Referential Example 4) (2S,4S)-4-Mercapto- 1 -methyl-2-[(3 S)-3-(N-methyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl)pyrrolidin-1-ylcarbonyl]pyrrolidine A solution of (2S,4S)-4-(4-methoxybenzylthio)-l-methyl-2pyrrolidinecarboxylic acid (0.49 g) in tetrahydrofuran (5 ml), N,Ndiisopropylethylamine (0.30 ml) and pivaloyl chloride (0.21 ml) were added in an ice bath. The resulting mixture was stirred at 0°C for 10 minutes. To the reaction mixture, a solution of a mixture of the compound (0.46 g) obtained in Referential Example 3-(2) and N,N-diisopropylethylamine (0.30 ml) in acetonitrile (10 ml) was added and the resulting mixture was stirred at 0°C for 30 minutes. Saturated saline solution was poured into the reaction mixture to terminate the reaction, followed by extraction three times with ethyl acetate. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol whereby (2S,4S)-4-(4methoxybenzylthio)-1 -methyl-2-[(3 S)-3-(N-methyl-N-(4nitrobenzyloxycarbonyl)aminomethyl)pyrrolidin-1-ylcarbonyl]pyrrolidine (0.76 g) was obtained.

Infrared absorption spectrum (Liquid film) v max cm': 1734,1705,1646,1609,1585,1513,1440,1403,1373,1346,1301,1248,1212,1191,1149, 1107.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 8 ppm: 1.55- 2.18(2H,m), 2.35,2.37(3H,sx2), 2.41-2.70(3H,m), 3.04,3.06(3H,sx2), 3.10- 3.88(10H,m), 3.75(2H,s), 3.85(3H,s), 5.27(2H,s), 6.89(2H,d,J=8.6Hz), 7.27(2H,d,J=7.9Hz), 7.57(2H,d,J=8.6Hz), 8.28(2H,d,J=7.9Hz).

To a solution of the compound (0.64 g) obtained in Referential Example 4in a mixture of anisole (0.65 ml) and trifluoroacetic acid (6.5 ml), trifluoromethanesulfonic acid (0.26 ml) was added while stirring in an ice bath, followed by stirring at room temperature for one hour. The reaction mixture was treated in a similar manner to that described in Referential Example whereby the title compound (0.54 g) was obtained.

Doc: 97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English translation/07.10.98 (Referential Example (2S,4S)-4-Mercapto-2-[(3R)-3-(N-methyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl)pyrrolidin- 1-ylcarbonyl]- 1-(4-nitrobenzyloxycarbonyl)pyrrolidine To a solution of the compound (1.17 which had been obtained in Referential Example in tetrahydrofuran (12 ml), triethylamine (0.89 ml) and methanesulfonyl chloride (0.50 ml) were added successively in an ice bath. The resulting mixture was stirred for one hour. Saturated saline solution was poured into the reaction mixture to terminate the reaction, followed by extraction three times with ethyl acetate. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure.

To the residue (1.66 a 40% methylamine-methanol solution (25 ml) was added and the resulting mixture was heated at 100 0 C for 4 hours in a pressure bottle.

After the temperature of the reaction mixture was allowed to lower to room temperature, the mixture was concentrated by evaporation under reduced pressure. To a solution of the residue (1.89 g) in acetonitrile (20 ml), N,N-diisopropylethylamine (1.84 ml) and 4nitrobenzyl chloroformate (2.28 g) were added in an ice bath and the resulting mixture was stirred at 0°C for 2 hours. The reaction mixture was treated in a similar manner to that described in Referential Example whereby (3S)-l-tert-butoxycarbonyl-3-[Nmethyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl]pyrolidine (1.92 g) was obtained.

Optical rotation: [a]D 2 5 +7.20 CHC13).

Infrared absorption spectrum (Liquid film) v max cm' 1 1696,1608,1524,1480,1455,1404,1366,1347,1293,1255,1211,1191,1170,1152,1125.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 8 ppm: 1.45(9H,s), 1.56-1.68(1H,m), 1.89-2.01(1H,m), 2.45-2.55(1H,m), 2.98(3H,s), 2.98-3.10(1H,m), 3.27-3.57(5H,m), 5.22(2H,s), 7.51(2H,d,J=8.6Hz), 8.23(2H,d,J=8.6Hz).

To a solution of the compound (0.84 g) obtained in Referential Example in methylene chloride (8 ml), trifluoroacetic acid (1.6 ml) was added in an ice bath.

The resulting mixture was stirred at room temperature for 2 hours. The reaction mixture was treated in a similar manner to that described in Referential Example whereby Doc: 97162p2.doc P79569/FP-9716(PCT)tsa-iglEnglish translation/07. 10.98 (3R)-3 ethyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl]pyrrolidine (0.58 g) was obtained.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 6 ppm: 1.30- 1.51(1H,m), 1.81-1.97(Hm), 2.20-2.69(3H,m), 2.85-3.06(6H,m), 3 .20-3.35(2Hm), 5.22(2H,s), 7.51(2H,d,J=8.6Hz), 8.22(2H,d,J=8.6Hz).

To a solution of (2S,4S)-4-(4-methoxybenzylthio)-1-(4nitrobenzyloxycarbonyl)-2-pyrrolidinecarboxylic acid (0.92 g) in tetrahydrofuran ml), N,N-diisopropylethylamine (0.36 ml) and pivaloyl chloride (0.25 ml) were added in an ice bath, followed by stirring at 0 0 C for 10 minutes. To the reaction mixture, a solution of a mixture of the compound (0.55 g) obtained in Referential Example 5-(2) and N,N-diisopropylethylamine (0.36 ml) in acetonitrile (10 ml) was added and the resulting mixture was stirred at 0 0 C for 30 minutes. The reaction mixture was treated in a similar manner to that described in Referential Example whereby (2S,4S)-4-(4methoxybenzylthio)-2-[(3R)-3-(N-methyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl)pyrrolidin- 1 -ylcarbonyl]- 1 4 -nitrobenzyloxycarbonyl)pyrrolidine (1.00 g) was obtained.

Infrared absorption spectrum (KBr) v max cm': 1706,1654,1608,1585,1520,1438,1403,1346,1299,1249,1210,1194,1175,1148,1109.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 6 ppm: 1.50- 2.68(5H,m), 2.92-4.08(9Hm), 2.97(3Hs), 3.73(2Hs), 3.79,3.80(3H,sx2), 4.30- 4.46(1H,m), 4.97-5.35(4Hm), 6.85(2Hd,J=8.6Hz), 7.24(2H,d,J=8.6Hz), 7.46(2H,d,J=8.6Hz), 7.51(2H,d,J=8.6Hz), 8.23(4H,d,J=8.6Hz).

To a solution of the compound (0.99 g) obtained in Referential Example in a mixture of anisole (1.0 ml) and trifluoroacetic acid (10 ml), trifluoromethanesulfonic acid (0.30 ml) was added in an ice bath. The resulting mixture was stirred at room temperature for one hour. The reaction mixture was treated in a similar manner to that described in Referential Example whereby the title compound (0.90 g) was obtained.

Doc: 97162p2.doc P 7 9569/FP-9716(PCT)/tsa-ig/English translation/07.10.98 (Referential Example 6) (2S,4S)-4-Mercapto- 1 -methyl-2-[(3R)-3-(N-methyl-N-(4-nitrobenzyloxy carbonyl)aminomethyl)pyrrolidin- 1 -ylcarbonyl]pyrrolidine To a solution of (2S,4S)-4-(4-methoxybenzylthio)- 1 -methyl-2pyrrolidinecarboxylic acid (0.50 g) in tetrahydrofuran (5 ml), N,Ndiisopropylethylamine (0.31 ml) and pivaloyl chloride (0.22 ml) were added in an ice bath. The resulting mixture was stirred at 0 0 C for 10 minutes. To the reaction mixture, a solution of a mixture of the compound (0.47 g) obtained in Referential Example 5-(2) and N,N-diisopropylethylamine (0.31 ml) in acetonitrile (10 ml) was added, followed by stirring at 0 0 C for 30 minutes. The reaction mixture was treated in a similar manner to that described in Referential Example whereby (2S,4S)-4-(4methoxybenzylthio)- 1 -methyl-2-[(3R)-3-(N-methyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl)pyrrolidin- 1 -ylcarbonyl]pyrrolidine (0.67 g) was obtained.

Infrared absorption spectrum (Liquid film) v max cm-: 1705,1647,1609,1585,1512,1440,1403,1346,1301,1248,1212,1191,1149,1107 Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 6 ppm: 1.50- 2.12(2H,m), 2.29,2.32(3H,sx2), 2.37-2.66(3H,m), 3.00,3.02(3Hsx2), 3.00- 3.88(10OH,m), 3.72(2H,s), 3.81(3H,s), 5.23(2H,s), 6.85(2H,d,J=8.6Hz), 7.23(2H,d,J=8.6Hz), 7.53(2H,d,J=8.6Hz), 8.24(2H,d,J=8.6Hz).

To a solution of the compound (0.59 which had been obtained in Referential Example in a mixture of anisole (0.6 ml) and trifluoroacetic acid ml), trifluoromethanesulfonic acid (0.24 ml) was added while stirring in an ice bath.

The resulting mixture was stirred at room temperature for one hour. The reaction mixture was treated in a similar manner to that described in Referential Example whereby the title compound (0.53 g) was obtained.

Doc: 97162p2.doc P79569/FP-971 6(PCT)/tsa-ig/English translation/07.1 O.98 (Referential Example 7) (2S,4S)-4-Mercapto-l-(4-nitrobenzyloxycarbonyl)-2-[(3R)-3-(4-nitrobenzyloxycarbonylguanidinomethyl)pyrrolidin- 1 -ylcarbonyl]pyrrolidine To a solution of the compound (1.50 g) obtained in Referential Example 1in acetonitrile (15 ml), triphenylphosphine (1.83 g) was added and the resulting mixture was refluxed for one hour. To the reaction mixture, sodium sulfate hydrate (2.24 g) was added, followed by reflux for a further one hour. After the temperature was allowed to lower to room temperature, the insoluble material was filtered off. The filtrate was then concentrated by evaporation under reduced pressure. Diethyl ether was poured into the residue and insoluble material so precipitated was filtered off. The filtrate was concentrated by evaporation under reduced pressure.

To a solution of the residue (1.90 g) in ethanol (20 ml), 1H-pyrazole-1carboxamidine hydrochloride (1.02 g) was added and the resulting mixture was refluxed for 3 hours. After the temperature of the reaction mixture was allowed to lower to room temperature, the mixture was concentrated by evaporation under reduced pressure. The residue was washed with diethyl ether.

To a solution of the residue (2.86 g) in a tetrahydrofuran (25 ml) water (25 ml) mixture, 4-nitrobenzyl chloroformate (3.57 g) and a 1N aqueous sodium hydroxide solution (33 ml) were added successively in an ice bath, followed by stirring at 0°C for minutes. The reaction mixture was diluted with water, followed by extraction three times with methylene chloride. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure. The residue so obtained was purified by chromatography through a silica gel column (ethyl acetate/methanol whereby 1-tert-butoxycarbonyl-3-(4-nitrobenzyloxycarbonylguanidinomethyl)pyrrolidine (1.29 g) was obtained.

Infrared absorption spectrum (KBr) v max cm'': 3393,3327,1693,1652,1641,1604,1523,1408,1347,1289.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 8 ppm: 1.44(9H,s), 1.68-1.52(1H,m), 1.95-2.04(1H,m), 2.36-2.48(1H,m), 3.03(1H,dd,J=1 1.2,7.2Hz), 3.15- 3.54(5H,m), 5.19(2H,s), 6.72(2H,br.s), 7.54(2H,d,J=8.6Hz), 8.20(2H,d,J=8.6Hz).

Doc: 97162p2.doc P79569/FP-9716(PCTytsa-ig/English translation/07.10.98 To a solution of the compound (1.23 g) obtained in Referential Example in methylene chloride (13 ml), trifluoroacetic acid (2.2 ml) was added in an ice bath. The resulting mixture was stirred at room temperature for 5 hours. The reaction mixture was treated in a similar manner to that described in Referential Example whereby 4 -nitrobenzyloxycarbonylguanidinomethyl)pyrrolidine (0.72 g) was obtained.

To a solution of 2 S,4S)-4-(4-methoxybenzylthio)-1-(4-nitrobenzyloxycarbonyl)-2-pyrrolidinecarboxylic acid (1.05 g) in tetrahydrofuran (10 ml), N,Ndiisopropylethylamine (0.41 ml) and pivaloyl chloride (0.29 ml) were added in an ice bath. The resulting mixture was stirred at 0°C for 10 minutes. To the reaction mixture, a solution of a mixture of the compound (0.72 g) obtained in Referential Example 7-(2) and N,N-diisopropylethylamine (0.41 ml) in acetonitrile (10 ml) was added, followed by stirring at 0°C for 30 minutes. Into the reaction mixture, saturated saline solution was poured to terminate the reaction, followed by extraction three times with ethyl acetate. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure.

The residue so obtained was purified by chromatography through a silica gel column (ethyl acetate/methanol whereby (2S,4S)-4-(4-methoxybenzylthio)-1-(4nitrobenzyloxycarbonyl)-2-[(3R)-3-(4-nitrobenzyloxycarbonylguanidinomethyl)pyrrolidin-l-ylcarbonyl]pyrrolidine (0.93 g) was obtained.

Infrared absorption spectrum (KBr) v max cm- 1707,1651,1609,1521,1432,1405,1346,1319,1287,1251,1207,1174,1110.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 8 ppm: 1.55- 2.71(5H,m), 2.96-4.40(10 3.72,3.74(2H,sx2), 3.79(3H,s), 4.97-5.38(4H,m), 6.81- 6.88(2H,m), 7.20-7.25(2Hm), 7.39-7.56(4H,m), 7 .99-8.25(4H,m).

To a solution of the compound (0.88 g) obtained in Referential Example 7in a mixture of anisole (1.4 ml) and trifluoroacetic acid (14 ml) trifluoromethanesulfonic acid (0.31 ml) was added while stirring in an ice bath. The resulting mixture was stirred at room temperature for one hour. The reaction mixture was treated in a similar manner to that described in Referential Example whereby the title compound (1.28 g) was obtained.

Doc: 97162p2.doc P79569/FP-9716(PC)/tsa-iglEnglish translation/0l. 10.98 (Referential Example 8) (2S,4S)-4-Mercapto- 1 -(4-nitrobenzyloxycarbonyl)-2-[(3R)-3-(4nitrobenzyloxycarbonylacetimidoylaminomethyl)pyrrolidin- -ylcarbonyl]pyrrolidine To a solution of the compound (1.00 g) obtained in Referential Example 1in acetonitrile (10 ml), triphenylphosphine (1.22 g) was added. The resulting mixture was refluxed for one hour. Sodium sulfate decahydrate (1.50 g) was added to the reaction mixture, followed by reflux for further one hour. After the temperature was allowed to rise to room temperature, insoluble material was filtered off. The filtrate was concentrated by evaporation under reduced pressure. Diethyl ether was poured into the residue. Insoluble material so precipitated was filtered off and the filtrate was concentrated by evaporation under reduced pressure.

To a solution of the residue (1.27 g) in acetonitrile (25 ml), a 4N hydrogen chloride ethyl acetate solution (1.1 ml) was added in an ice bath, followed by stirring at 0°C for 15 minutes. To the reaction mixture, N-(4nitrobenzyloxycarbonyl)acetamidine (1.15 g) was added in an ice bath and the resulting mixture was stirred at 50 0 C for 1.5 hours. After the temperature of the reaction mixture was allowed to lower to room temperature, it was diluted with ethyl acetate. The diluted solution was washed with a saturated aqueous solution of sodium bicarbonate and saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure. The residue so obtained was purified by chromatography through a silica gel column, whereby (3R)-1-tert-butoxycarbonyl-3-[N- 4 -nitrobenzyloxycarbonyl)acetimidoylaminomethyl)pyrrolidine (1.66 g) was obtained.

Infrared absorption spectrum (Liquid film) v max cm" 1 3313,3112,1687,1564,1523,1411,1346,1220.

Nuclear magnetic resonance spectrum (270 MHz, CDC1 3 8 ppm: 1.46(9H,s), 1.58-1.72(1H,m), 1.95-2.12(1H,m), 2.16,2.25(3H,sx2), 2.39-2.53(1H,m), 3.00- 3.11(1H,m), 3.28-3.62(5H,m), 5.20,5.23(2H,sx2), 7.57(2H,d,J=8.6Hz), 8.21(2H,d,J=8.6Hz).

To a solution of the compound (1.66 g) obtained in Referential Example 8in methylene chloride (17 ml), trifluoroacetic acid (2.9 ml) was added in an ice bath.

The resulting mixture was stirred at room temperature for 2 hours. The reaction mixture Doc: 97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English translation/07.10.98 was treated in a similar manner to that described in Referential Example whereby 4 -nitrobenzyloxycarbonyl)acetimidoylaminomethyl]pyrrolidine (0.81 g) was obtained.

To a solution of (2S,4S)-4-(4-methoxybenzylthio)-l-( 4 -nitrobenzyloxycarbonyl)-2-pyrrolidinecarboxic acid (1.18 g) in tetrahydrofuran (12 ml), N,Ndiisopropylethylamine (0.46 ml) and pivaloyl chloride (0.33 ml) were added in an ice bath. The resulting mixture was stirred at 0°C for 10 minutes. To the reaction mixture, a solution of a mixture of the compound (0.81 obtained in Referential Example 8-(2) and N,N-diisopropylethylamine (0.46 ml) in acetonitrile (10 ml) was added, followed by stirring at 0°C for 30 minutes. Saturated saline solution was poured into the reaction mixture to terminate the reaction, followed by extraction three times with ethyl acetate.

The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure. The residue so obtained was purified by chromatography through a silica gel column (ethyl acetate/methanol 95/5 whereby (2S,4S)-4-(4-methoxybenzylthio)-1-(4nitrobenzyloxycarbonyl)-2-[(3R)-3-(N-(4-nitrobenzyloxycarbonyl)acetimidoylaminomethyl)pyrrolidin- 1-ylcarbonyl]pyrrolidine (1.05 g) was obtained.

Infrared absorption spectrum (KBr) v max cm- 1 3300,3113,3080,1709,1651,1608,1585,1564,1520,1439,1404,1346,1301,1241,1213, 1176,1110.

Nuclear magnetic resonance spectrum (270 MHz, CDC13) 6 ppm: 1.49- 2.77(8H,m), 3.03-4.12(10H,m), 3.79(2H,s), 3.84(3H,s), 4.30-4.50(1H,m), 5.04- 5.40(4H,m), 6.90(2H,d,J=8.6Hz), 7.29(2H,d,J=8.6Hz), 7.43-7.65(4H,m), 8.16- 8.32(4H,m).

To a solution of the compound (1.02 g) obtained in Referential Example 8in a mixture of anisole (1.5 ml) and trifluoroacetic acid (15 ml), trifluoromethanesulfonic acid (0.36 ml) was added while stirring in an ice bath. The resulting mixture was stirred at room temperature for one hour. The reaction mixture was treated in a similar manner to that described in Referential Example whereby the title compound (1.35 g) was obtained.

Doc: 97162p2.doc P79569/FP-9716PCj'Iytsa-ig/English translation/07.10.98 (Referential Example 9) S)-3-(N-cyclopropyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl)pyrrolidin-1-ylcarbonyl]-4-mercapto-1-(4-nitrobenzyloxycarbonyl]pyrrolidine.

To a solution of the compound (1.00 g) obtained in Referential Example 1in tetrahydrofuran (10 ml), triethylamine (0.83 ml) and methanesulfonyl chloride (0.46 ml) were successively added in an ice bath. The resulting mixture was stirred for one hour. Saturated saline solution was poured into the reaction mixture to terminate the reaction, followed by extraction three times with ethyl acetate. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure.

To a solution of the residue (1.53 g) in methanol (10 ml), cyclopropylamine ml) was added and the resulting mixture was heated at 100 0 C for 4 hours in a pressure bottle. After the temperature of the reaction mixture was allowed to lower to room temperature, the mixture was concentrated by evaporation under reduced pressure. To a solution of the residue (1.93 g) in acetonitrile (20 ml), N,N-diisopropylethylamine (0.95 ml) and 4-nitrobenzyl chloroformate (1.18 g) were added in an ice bath and the resulting mixture was stirred at room temperature for 2 hours. Saturated saline solution was poured into the reaction mixture to terminate the reaction, followed by extraction three times with ethyl acetate. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure. The residue so obtained was purified by chromatography through a silica gel column (n-hexane/ethyl acetate whereby (3S)-l-tertbutoxycarbonyl-3-[N-cyclopropyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl]pyrrolidine (0.95 g) was obtained.

Infrared absorption spectrum (Liquid film) v max cm' 1 1697,1607,1524,1494,1479,1453,1405,1366,1346,1288,1258,1210,1171,1141,1111.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 8 ppm: 0.63- 0.71(2H,m), 0.78-0.87(2H,m), 1.45(9H,s), 1.55-1.68(1H,m), 1.90-2.00(1H,m), 2.50- 2.67(2H,m), 2.96-3.09(lH,m), 3.18-3.59(5H,m), 5.23(2H,s), 7.53(2H,d,J=8.6Hz), 8.23(2H,d,J=8.6Hz).

Doc: 97162p2.doc P79569/FP-9716(PCT)/tsa-iglEnglish translation/07.1O.98 To a solution of the compound (0.77 g) obtained in Referential Example 9in methylene chloride (12 ml), trifluoroacetic acid (1.4 ml) was added in an ice bath, followed by stirring at room temperature for 3 hours. The reaction mixture was treated in a similar manner to that described in Referential Example whereby cyclopropyl-N-(4-nitrobenzyloxycarbonyl)aminomethyl]pyrrolidine (0.72 g) was obtained.

To a solution of (2S,4S)-4-(4-methoxybenzylthio)-1-(4nitrobenzyloxycarbonyl)-2-pyrrolidinecarboxylic acid (1.00 g) in tetrahydrofuran ml), N,N-diisopropylethylamine (0.39 ml) and pivaloyl chloride (0.28 ml) were added in an ice bath. The resulting mixture was stirred at 0°C for 10 minutes. To the reaction mixture, a solution of a mixture of the compound (0.72 obtained in Referential Example and N,N-diisopropylethylamine (0.39 ml) in acetonitrile (10 ml) was added, followed by stirring at 0°C for 30 minutes. Saturated saline solution was poured into the reaction mixture to terminate the reaction, followed by extraction three times with ethyl acetate. The combined organic layer was washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (ethyl acetate), whereby (2S,4S)-2-[(3S)-3-(N-cyclopropyl-N-(4nitrobenzyloxycarbonyl)aminomethyl)pyrolidin-1-ylcarbonyl]-4-(4methoxybenzylthio)-1-(4-nitrobenzyloxycarbonyl)pyrrolidine (0.93 g) was obtained.

Infrared absorption spectrum (KBr) v max cm- 1 1707,1655,1609,1585,1521,1440,1404,1346,1289,1250,1210,1176,1139,1110.

Nuclear magnetic resonance spectrum (270 MHz, CDC1 3 6 ppm: 0.50- 0.75(2H,m), 0.75-0.95(2H,m), 1.47-2.02(3H,m), 2.32-2.74(3H,m), 2.92-4.08(9H,m), 3.73(2H,s), 3.79,3.80(3H,sx2), 4.26-4.45(1H,m), 4.97-5.34(4H,m), 6.85(2H,d,J=8.6Hz), 7.23,7.25(2H,d2,J=8.6Hz), 7.46,7.53(4H,dx2,J=8.6Hz), 8.23(4H,d,J=8.6Hz).

To a solution of the compound (0.82 g) obtained in Referential Example 9in a mixture of anisole (0.8 ml) and trifluoroacetic acid (8 ml), trifluoromethanesulfonic acid (0.24 ml) was added while stirring in an ice bath, followed by stirring at room temperature for one hour. The reaction mixture was treated Doc: 97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English branslation/07.10.98 in a similar manner to that described in Referential Example whereby the title compound (0.77 g) was obtained.

(Referential Example (2S,4S)-4-Mercapto-l-(4-nitrobenzyloxycarbonyl)-2-[(3S)-3-(4-nitrobenzyloxycarbonylacetimidoylaminomethyl)pyrrolidin- 1 -ylcarbonyl]pyrrolidine.

To a solution of the compound (1.00 g) obtained in Referential Example 2in acetonitrile (10 ml), triphenylphosphine (1.22 g) was added and the resulting mixture was refluxed for one hour. To the reaction mixture, sodium sulfate decahydrate (1.50 g) was added, followed by reflux for further one hour. After the temperature of the reaction mixture was allowed to lower to room temperature, insoluble material was filtered off. The filtrate was then concentrated by evaporation under reduced pressure.

Into the residue, diethyl ether was poured and insoluble material so precipitated was filtered off. The filtrate was then concentrated by evaporation under reduced pressure.

To a solution of the residue (1.33 g) in acetonitrile (25 ml), a 4N hydrogen chloride ethyl acetate solution (1.1 ml) was added in an ice bath and the resulting mixture was stirred at 0°C for 15 minutes. To the reaction mixture, N-(4nitrobenzyloxycarbonyl)acetamidine (1.15 g) was added in an ice bath. The resulting mixture was stirred at 50 0 C for 1.5 hours. The reaction mixture was treated in a similar manner to that described in Referential Example whereby (3S)-1-tertbutoxycarbonyl-3-[(N- 4 -nitrobenzyloxycarbonyl)acetimidoylaminomethyl]pyrrolidine (1.58 g) was obtained.

Infrared absorption spectrum (Liquid film) v max cm' 1 3314,3112,1690,1565,1523,1409,1346,1221.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 S ppm: 1.46(9H,s), 1.58-1.72(1H,m), 1.95-2.12(1H,m), 2.16,2.25(3H,sx2), 2.39-2.53(1H,m), 3.00- 3.11(1H,m), 3.28(5H,m), 5.20,5.23(2H,sx2), 7.57(2H,d,J=8.6Hz), 8.21(2H,d,J=8.6Hz).

To a solution of the compound (1.58 g) obtained in Referential Example in methylene chloride (15 ml), trifluoroacetic acid (2.9 ml) was added in an ice bath.

The resulting mixture was stirred at room temperature for 2 hours. The reaction mixture Doc: 97162p2.doc P79569/FP-9716(PClytsa-igEnglish translation/07. 10.98 was treated in a similar manner to that described in Referential Example whereby (3R)-3-[N-(4-nitrobenzyloxycarbonyl)acetimidoylaminomethyl]pyrrolidine (0.97 g) was obtained.

To a solution of (2S,4S)-4-(4-methoxybenzylthio)-1-(4-nitrobenzyloxycarbonyl)-2-pyrrolidinecarboxylic acid (1.06 g) in tetrahydrofuran (10 mi), N,Ndiisopropylethylamine (0.41 ml) and pivaloyl chloride (0.29 ml) were added in an ice bath. The resulting mixture was stirred at 0 0 C for 10 minutes. To the reaction mixture, a solution of a mixture of the compound (0.76 g) obtained in Referential Example and N,N-diisopropylethylamine (0.41 ml) in acetonitrile (10 ml) was added, followed by stirring at 0 0 C for 30 minutes. The reaction mixture was treated in a similar manner to that described in Referential Example whereby (2S,4S)-4-(4methoxybenzylthio)- 1 -(4-nitrobenzyloxycarbonyl)-2-[(3S)-3-(N-(4-nitrobenzyloxycarbonyl)acetimidoylaminomethyl)pyrrolidin- 1 -ylcarbonyl]pyrrolidine (1.00 g) was obtained.

Infrared absorption spectrum (KBr) v max cm-1: 3296,3113,3080,1708,1680,1652,1608,1584,1564,1521,1432,1405,1346,1301,1239, 1216,1176,1110.

Nuclear magnetic resonance spectrum (270 MHz, CDCl 3 8 ppm: 1.60- 1.80(8H,m), 3.02-4.10(9Hm), 3.78(2Hs), 3.83(3,s), 4.32-4.52(1Hm), 5.02- 5.38(4H,m), 6.90(2H,d,J=8.6Hz), 7.28(2Hd,J=8.6Hz), 7.45-7.69(4H,m), 8.20- 8.34(4H,m).

To a solution of the compound (1.00 g) obtained in Referential Example in a mixture of anisole (1 ml) and trifluoroacetic acid (10 ml), trifluoromethanesulfonic acid (0.20 ml) was added while stirring in an ice bath. The resulting mixture was stirred at room temperature for two hours. The reaction mixture was treated in a similar manner to that described in Referential Example whereby the title compound (0.93 g) was obtained.

Doc: 97162p2.doc P79569/FP-9716(PCTytsa-igEnglish translation/07.1O.98 (Referential Example 11) (2S,4S)-4-Mercapto-l-(4-nitrobenzyloxycarbonyl)-2-[(3S)-3-(4nitrobenzyloxycarbonylguanidinomethyl)pyrolidin- -ylcarbonyl]pyrrolidine To a solution of the compound (1.01 g) obtained in Referential Example 2in anhydrous acetonitrile (10 ml), triphenylphosphine (1.23 g) was added and the resulting mixture was refluxed for one hour. To the reaction mixture, 1.51 g of sodium sulfate decahydrate were added, followed by reflux for further one hour. After cooling to room temperature, the reaction mixture was filtered. The filtrate was then concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol whereby (3S)-3-aminomethyl-l-tert-butoxycarbonylpyrrolidine (840 mg) was obtained. The product completely coincided with the compound obtained in Referential Example 1-(9) in HPLC and NMR data.

To a solution of the compound (840 mg) obtained in in ethanol (14 ml), 1H-pyrazole-l-carboxamidine hydrochloride (686 mg) was added and the resulting mixture was refluxed for 3 hours. After the temperature of the reaction mixture was allowed to lower to room temperature, the mixture was concentrated by evaporation under reduced pressure. The residue was washed with diisopropyl ether.

To a solution of the residue (1.50 g) in a tetrahydrofuran (20 ml) water (20 ml) mixture, 4-nitrobenzyl chloroformate (2.41 g) and a IN aqueous sodium hydroxide solution (22 ml) were successively added and the resulting mixture was stirred at 0 C for 30 minutes. The reaction mixture was diluted with water, followed by extraction three times with ethyl acetate. The combined organic layers were washed with saturated saline solution, dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure. The residue so obtained was purified by chromatography through a silica gel column (ethyl acetate/methanol 25/1), whereby (3S)-1-tertbutoxycarbonyl-3-(4-nitrobenzyloxycarbonylguanidinomethyl)pyrrolidine (746 mg) was obtained.

Nuclear magnetic resonance spectrum (270 MHz, CDC13) 8 ppm: 1.44(9H,s), 1.68-1.52(1H,m), 1.95-2.04(1H,m), 2.36-2.48(1H,m), 3.03(1H,dd,J=l 1.1,7.1Hz), 3.15- 3.54(5H,m), 5.19(2H,s), 6.70(1H,br.s), 7.54(2H,d,J=8.6Hz), 8.20(2H,d,J=8.6Hz).

Doc: 97162p2.doc P79569/FP-9716(PCT)tsa-ig/English translation/07. 10.98 To a solution of the compound (701 mg) obtained in in ethyl acetate (7 ml), a 4N hydrochloric acid ethyl acetate solution (3.3 ml) was added and the resulting mixture was stirred at room temperature for one hour. The reaction mixture was concentrated. The residue was then washed with ethyl ether, whereby nitrobenzyloxycarbonylguanidinomethyl)pyrrolidine hydrochloride (647 mg) was obtained.

To (2S,4S)-4-(4-methoxybenzylthio)-1 4 -nitrobenzyloxycarbonyl)-2pyrrolidinecarboxylic acid (947 ml) in N,N-dimethylformamide (10 ml), N,Ndiisopropylethylamine (0.369 ml), the compound obtained in (647 mg), 1-ethyl-3-(3dimethylaminopropyl)carbodiimide hydrochloride (406 mg) and 1hydroxybenzotriazole (286 mg) were added and the resulting mixture was stirred overnight at room temperature. Ethyl acetate was added to the reaction mixture. The resulting mixture was washed with water and saturated saline solution, dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure.

The residue so obtained was separated by chromatography through a silica gel column (ethyl acetate/methanol 12/1), whereby (2S,4S)-4-(4-methoxybenzylthio)-1-(4nitrobenzyloxycarbonyl)-2-[(3

S)-

3 4 -nitrobenzyloxycarbonylguanidinomethyl)pyrolidin- -ylcarbonyl]pyrrolidine (1.13 g) was obtained.

Infrared absorption spectrum (KBr) v max 1708,1520,1251,1032, 739.

Nuclear magnetic resonance spectrum (270 MHz, CDC13) 8 ppm: 1.5-2.7(5H,m), 2.9-3.6(10H,m), 3.71,3.73(2H,sx2), 3.77(3H,s), 3.80-3.90(1H,m), 4.95-5.25(4H,m), 6.87(2H,d,J=8.45Hz), 7.25-7.28(2H,m), 7.45-7.65(4H,m), 8.15-8.3(4H,m).

To a solution of the compound (1.10 g) obtained in in a mixture of anisole (1.65 ml) and trifluoroacetic acid (8.0 ml), trifluoromethanesulfonic acid (0.26 ml) was added while stirring in an ice bath. The resulting mixture was stirred at the same temperature for 30 minutes. The reaction mixture was treated as in Referential Example whereby the title compound (926 mg) was obtained.

Doc: 97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English tb-nslation/07.10.98 (Referential Example 12) (2S,4S)-4-Mercapto-l-(4-nitrobenzyloxycarbonyl)-2-[(3R)-3-(pyrrolidin-1ylmethyl)pyrrolidin-1-ylcarbonyl]pyrrolidine To a solution of (3S)-3-(pyrrolidin-1-ylmethyl)pyrrolidine hydrochloride (367 mg), which had been prepared from 3 R)-1-tert-butoxycarbonyl-3hydroxymethylpyrrolidine, in anhydrous dimethylformamide (10 ml), (2S,4S)-4-(4methoxybenzylthio)-1-( 4 -nitrobenzyloxycarbonyl)-2-pyrrolidinecarboxylic acid (804 mg), diisopropylethylamine (1.16 ml), 1-hydroxybenzotriazole (243 mg) and 1-ethyl-3- (3-dimethylaminopropyl)carbodiimide hydrochloride (483 mg) were added. The resulting mixture was stirred at room temperature for 10 hours. The solvent was then distilled off under reduced pressure. The residue was dissolved in ethyl acetate. The resulting solution was washed with a saturated aqueous solution of sodium bicarbonate.

The organic layer was dried over anhydrous sodium sulfate and then concentrated by evaporation under reduced pressure. The residue was purified by chromatography through a silica gel column (ethyl acetate/methanol 5/1 whereby (2S,4S)-4-(4methoxybenzylthio)-2-[(3R)-3-(pyrrolidin- 1-ylmethyl)pyrrolidin- 1 -ylcarbonyl]- 1 nitrobenzyloxycarbonyl)pyrrolidine (362 mg) was obtained.

Nuclear magnetic resonance spectrum (270 MHz, CDC13) 6 ppm: 1.00-2.00(3H, 2.00-3.00(2H, 3.00-4.20(17H, 3.73(2H, 3.79(3H, 4.32-4.46(1H, m), 4.90-5.40(2H, 6.85(2H, d, J=8.5 Hz), 7.24(2H, d, J=8.5 Hz), 7.46(2H, d, J=8.6 Hz), 8.23(2H, d, J=8.6 Hz).

To a solution of the compound (362 mg) obtained in in a mixture of anisole (0.68 ml) and trifluoroacetic acid (3.4 ml), trifluoromethanesulfonic acid (0.14 ml) was added while stirring in an ice bath. The resulting mixture was stirred at the same temperature for 30 minutes. Trifluoroacetic acid was distilled off under reduced pressure. The residue was washed with hexane and ether, dried under reduced pressure, extracted with ethyl acetate after addition of a saturated aqueous solution of sodium bicarbonate and then washed with saturated saline solution. The organic layer was dried over anhydrous sodium sulfate and concentrated by evaporation under reduced pressure, whereby the title compound (287 mg) was obtained as a powder.

Infrared absorption spectrum (KBr) v max cm': 1697, 1525, 1246.

Doc: 97162p2.doc P79569/FP-9716(PCTytsa-ig/Engish translation/07. 10.98 (Referential Example 13) 2 S, 4

S)-

2 3 R)-3 -(N-2-Hydroxyethyl N-4nit-obenzyloxycarbonylam inom ethyl)pyrrol idin- I -yl carbonyl]-4-m ercapto- 1 4 -nitrob enzyloxycarbonyl)-pyrroli dine (3 S)- 3 2 -Hydroxyethyl-N-4-nitrobenzyloxycarbonylaminomethyl)pyrrolidine hydrochloride (197 mg) was treated in a similar manner to that described in Referential Example 12-(1) and to afford the title compound (214 mg) as a powder.

Nuclear magnetic resonance spectrum (270 MI~z, CDCI 3 8 PPM: 1.50-2.20(2K71 in), 2.50-2.90(2, in), 2.90-4.10(12K in), 4.15-4.30(2K in), 4.40-4.55(1H, in), 5.20- 5.30(4H, in), 7.50-7.60(4H, in), 8.19-8.28(4H, in).

(Referential Example 14) (2 S)-3 -(N-carbamoylmethyl-N-4-nitrobenzyloxycarbonylaminomethyl)pyrrolidin- 1 -yl carbonyl]-4-m ercapto- 1 4 -nitrobenzyloxycarbonyl)pyrro i dine (3 S)- 3 -(N-Carbamoylmethyl-N-4-nitrobenzyloxycarbonylaminomethyl).

pyrrolidine hydrochloride (261 mg) was treated in a similar manner to that described in Referential Example 12-(1) and to give the title compound (368 mg) as a powder.

Infrared absorption spectrum (KBr) v max cm": 3371, 2948, 2879, 1706, 1645, 1608, 1522, 1347, 1259.

Nuclear magnetic resonance spectrum (400 MHz, DMSO-d 6 5 PPM: 1.40- 2.05(2H, in), 2.30-2.75(3H, mn), 2.90-3.60(5H, in), 3.60-4.95(4K in), 4.30-4.60(I, in), 5.00-5.30(4H, in), 7.40-7.70(4H, in), 8.10-8.30(4H, in).

(Referential Example (2S,4S)-4-Mercapto- I -(4-nitrobenzyloxycarbonyl)-2-[(3 nitrobenzyloxycarbonylamino)ethyl-N-4nitrobenzyloxycarbonylaminom ethyl] pyrrolidin- l-ylcarbonyllpyrrolidine (3 S)- 3 -[-2-(4-nitrobenzyloxycarbonylamino)ethyl..N-4 nitrobenzyloxycarbonylaminomethyl]pyrrolidine hydrochloride (195 mg) was treated in P79569/FP-9716(PCT)/tsa-igEnglish translatonO7. 10.98 a similar manner to that described in Referential Example 12-(1) and to give the title compound (248 mg) as a powder.

Infrared absorption spectrum (KBr) v max cm-i: 1705, 1648, 1607, 1521, 1437, 1347.

Nuclear magnetic resonance spectrum (270 Mfhz, CDC1 3

CD

3 OD) 5 ppm: 1.50-2.10(3H1, in), 2.60-2.80(2K1 mn), 3.00-3.70(l111, in), 3.80-4.00(1H, in), 4.00- 4.20(111, in), 4.40-4.60(111, in), 5.15-5.30(6H1, in), 7.45-7.60(6K1 in), 8.10-8.30(6K1 in).

(Referential Example 16) (2 S,4S)-2-r(3 S)-3 -(N-2-Diinethylaminoethyl-N-4-nitrobenzyloxycarbonyl.

aininoinethyl)pyrrolidin- I -ylcarbonyl]-4-inercapto-l1-(4-nitrobenzyloxycarbonyl)pyrrolidine (3 S)- 3 -(N-2-Dimethylaminoethyl-N-4-nitrob enzyloxycarbonylam inon ethyl)-.

pyrrolidine dihydrochloride (302 mg) was treated in a similar manner to that described in Referential Example 12-(1) and to afford the title compound (461 ing) as a powder.

(Referential Example 17) (2 S,4S)-4-Mercapto-2-[(3R)-3 -IN-2-(N-inethyl-N-4-nitrobenzyloxycarbonylamino)ethyl-N-methylaininomethyl]pyrrolidin- 1 -ylcarbonyll- 1 -(4-nitrobenzyloxycarbonyl)pyrrolidine (3 S)- 3 -IIN-2-(N-Methyl-N-4-nitrobenzyloxycarbonylanino)ethyl.N inethylaminoinethyl]pyrrolidine dihydrochloride (580 ing) was treated in a similar manner to that described in Referential Example 12-(l) and to give the title compound (628 mg) as a powder.

Infrared absorption spectrum (KBr) v max 1699, 1645, 1523, 1442, 1347.

Nuclear magnetic resonance spectrum (270 MHz, DMSO-d 6 5 PPM: 1.40- 2.00(3H1, in), 2.00-4.10(21H1, in), 4.30-4.60(111, mn), 5.00-5.30(4K1 in), 7.51-7.65(41L, in), 8.21-8.28(4K1 in).

97162p2.doe 971622.docP79569/FP-97l6(PCTytsa-ig/English traslationl07. 10.98 (Referential Example 18) (2 S,4 S)-3 -(N-2-Fluoroethyl-N-4-nitrob enzyloxycarbonylaminom ethyl pyrrolidin- 1 -ylcarbonyl]-4-mercapto-l1-(4-nitrobenzyloxycarbonyl)pyrrolidine (3 S)-3-(N-2-Fluoroethyl-N-4-nitrobenzyloxycarbonylaminomethyl)pyrrolidine hydrochloride is treated in a similar manner to that described in Referential Example 12-(l) and to afford the title compound.

(Referential Example 19) (2 S,4 S)-4-Mercapto-2-[(3R)-3 -[1I -methyl-2,3 -bis(4-nitrobenzyloxycarbonyl)guanidinomethyllpyrrolidin- I -ylcarbonyl]- 1 -(4-nitrob enzyloxycarbonyl)pyrrol idine (3 S)-3 -[1I -Methyl-2,3 -bis(4-nitrobenzyloxycarbonyl)guanidinomethyl]pyrrolidine hydrochloride (1.20 g) was treated in a similar manner to that described in Referential Example 12-(1) and to give the title compound (996 mg).

Infrared absorption spectrum (KBr) v max cm'1: 1754, 1709, 1645, 1608, 1521, 1441, 1405, 1347.

Nuclear magnetic resonance spectrum (270 MHz, DMSO-d 6 5 PPM: 1.50- 2.00(2H1, in), 2.60-2.80(1H, in), 2.90-3.05(4H, in), 3.10-3.60(7K1 mn), 3.65-3.80(2R1 in), 3.80-4.05(111, in), 4.30-4.60(111, mn), 5.00-5.30(6H1, in), 7.45-7.70(6H1, in), 8.05- 8.30(6H, in).

(Referential Example (2S,4 S)-4-Mercapto-lI-(4-nitrobenzyloxycarbonyl)-2-[(3R)-3 nitrobenzyloxycarbonylacetimidoyl-N-methylaminomethyl)pyrrolidinylcarbonyl]pyrroli dine (3 S)- 3 -(N-4-Nitrobenzyloxycarbonylacetimidoyl-N-inethylaminom ethyl)pyrrolidine hydrochloride is treated in a similar manner to that described in Referential Example 12-(1) and to afford the title compound.

P79569/FP-9716(PCT/tsa-ig/English translationO7. 10.98 (Referential Example 21) (2 S,4S)-4-Mercapto- 1 -(4-nitrobenzyloxycarbonyl)-2-[(3R)-3 nitrobenzyloxycarbonylformimidoyl-N-methylaminomethyl)pyrrolidin- 1ylcarbonyl]pyrrolidine (3 S)-3 -(N-4-Nitrobenzyloxycarbonylformimidoyl-N-methylaminomethyl)pyrrolidine hydrochloride is treated in a similar manner to that described in Referential Example 12-(1) and to give the title compound.

(Referential Example 22) (2S,4S)-4-Mercapto- 1 -(4-nitrobenzyloxycarbonyl)-2-[(3 S)-3-(N-2,2,2-trifluoroethyl-N- 4-nitrob enzyloxycarbonylaminom ethyl)pyrrol idin- 1 -ylcarbonyllpyrrolidine (3 S)- 3 -(N-2,2,2-Trifluoroethyl-N-4-nitrob enzyloxycarbonylaminom ethyl)pyrrolidine hydrochloride is treated in a similar manner to that described in Referential Example 12-(1) and to afford the title compound.

(Referential Example 23) (2 S,4 S)-4-Mercapto- 1 -(4-nitrobenzyloxycarbonyl)-2-[(3R)-3 nitrob enzyloxycarbonylformimidoylaminomethyl)pyrro i din- 1 -ylcarbonyllpyrrolidine (3 S)-3 -(4-Nitrob enzyloxycarbonylformim idoylm ethylam inomethyl)pyrroli dine hydrochloride is treated in a similar manner to that described in Referential Example 12-(1) and to afford the title compound.

(Referential Example 24) S)-3 -B is(4-nitrobenzyloxycarbonyl)guanidinom ethyl] pyrrolidin- I1ylcarbonyl]-4-mercapto- 1 -methylpyrrolidine (3 S)- 3 -[2,3-bis(4-Nitrobenzyloxycarbonyl)guanidinomethyl]pyrrolidine is treated in a similar manner to that described in Referential Example and to give the title compound.

Doc: 97162p2.doc Doc: 7162p.dacP795691FP-9716(PCTytsa-igEnglish tanslationO7. 10.98 (Referential Example (2 S,4S)-4-Mercapto- 1-methyl-2-[(3 S)-3 -(4-nitrobenzyloxycarbonylacetimidoylaminomethyl)pyrrol idin- 1 -ylcarbonyllpyrrolidine (3 S)- 3 4 -Nitrob enzyloxycarbonylacetim idoylaminom ethyl)pyrroli dine is treated in a similar manner to that described in Referential Example and to afford the title compound.

(Referential Example 26) (2S,4 S)-4-Mercapto- 1 -methyl-2-[(3R)-3-(4-nitrobenzyloxycarbonylformimidoyl.

aminomethyl)pyrrolidin- 1 -ylcarbonyllpyrrolidine (3 S)- 3 4 -Nitrobenzyloxycarbonylformimidoylaminomethyl)pyrroli dine is treated in a similar manner to that described in Referential Example and to give the title compound.

(Referential Example 27) (2S,4 1 -Methyl-2,3 -bis(4-nitrobenzyloxycarbonyl)guanidinomethyl-4mercapto- 1 -methylpyrrolidin- 1 -ylcarbonyllpyrrolidine (3 1 -Methyl-2,3 -bis(4-Nitrobenzyloxycarbonyl)guanidinomethyl].

pyrrolidine is treated in a similar manner to that described in Referential Example 12and to afford the title compound.

(Referential Example 28) (2S,4 S)-4-Mercapto- 1 -methyl-2-[(3 S)-3-(N-4-nitrobenzyloxycarbonylacetimidoyl-Nmethylaminomethyl)pyrrolidin- 1 -ylcarbonyl]pyrrolidine (3 S)- 3 -(N-4-Nitrobenzyloxycarbonylacetimidoyl-N-.methylaminomethyl).

pyrrolidine is treated in a similar manner to that described in Referential Example 4-(1) and to afford the title compound.

Doc: 97162p2.doc Doc: 7l62p.docP79569/FP-9716(PCI)/tsa-ig/English translationO7.1O.98 (Referential Example 29) (2S,4S)-4-Mercapto-1-methyl-2-[(3R)-3-(N-4-nitrobenzyloxycarbonylformimidoyl-Nmethylaminomethyl)pyrrolidin- 1-ylcarbonyl]pyrrolidine (3 S)- 3 4 -Nitrobenzyloxycarbonylformimidoyl-N-methylaminomethyl)pyrrolidine is treated in a similar manner to that described in Referential Example 4-(1) and to give the title compound.

(Test 1) In vitro Antibacterial activity Antibacterial activity was measured by the agar plate dilution method, whereby the minimum growth inhibitory concentration (MIC: jpg/ml) against various pathogenic bacteria was determined. The test results on the antibacterial activity of the compounds of the invention of Examples 3 and 7 against Staphylococcus aureus 209P, Escherichia coli NIHJ and Pseudomonas aeruginosa No. 7 are shown in Table 2.

Incidentally, Compounds A and B used for comparison are compounds of Example 3 in Japanese Patent Application Kokai No. Hei 5-310740 [compounds represented by the formula of the present invention, wherein R 2 and R 3 represent hydrogen atoms]. Compound A has the R configuration at the binding position of the aminomethyl group. Compound B has the S configuration at the binding position of the aminomethyl group.

P79569/FP-9716(PCT)/tsa-ig/English translation/07.10.98 [Table 2] Minimum Growth Inhibitory Concentration (MIC: Rig/ml) Bacterial strain tested Staphylococcus aureus 209P Eseherichia Pseudomonas co/i NJHJ aeruginosa No. 7 Compound of Ex. 3 0.02 0.02 0.05 Compound of Ex. 7 0.02 0.02 0.05 Compound A 0.02 0.02 0.1 Compound B 0.02 0.02 0.1 The medium used for the measurement was Mueller-Hinton 11 agar (MHA, Becton Dikinson Microbiology Systems).

(Test 2) In vitro Antibacterial activit The antibacterial activities of the compounds of the invention of Examples 3 and 7 and meropenern against Staphylococcus aureus 209P, Escherichia coli NIHJ and Pseudomonas aeruginosa 3719 (strain resistant to meropenem) are shown in Table 3.

97162p2.doe P79569/FP-9716(PCI)/tsa-ig/English translation/07.10.98 [Table 3] Minimum Growth Inhibitory Concentration (MIC: ptg/ml) Bacterial strain tested Staphylococcus Escherichia Pseudomonas aureus 209P coli NIHJ aeruginosa 3719 Compound ofEx. 3 0.01 0.01 0.2 Compound of Ex. 7 <0.01 0.01 0.1 Meropenem 0.02 <0.01 6.2 The medium used for the measurement was Nutrient Agar Eiken (Eiken Chemicals Co., Ltd.).

(Test 3) In vivo Antibacterial activity (treatment for infections) The culture of the bacterial strain to be tested was inoculated intraperitoneally to groups of mice (SPF, DDY, male), each group consisting of 7 animals. A solution of the test compound was subcutaneously administered twice in total to the mice, once immediately and once four hours after the inoculation. From the survival ratio five days after infection, the 50% effective dose (EDso: mg/kg) was determined by the Probit method and the single dose of test compound was indicated.

The test results of the compound of Example 3, compound A and Meropenem against Pseudomonas aeruginosa 1008 are shown in Table 4. Incidentally, Compound A is as described above.

97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English translation/07.10.98 [Table 4] Treatment against infections (ED 5 o: mg/kg) Bacterial strain tested Pseudomonas aeruginosa 1008 Compound of Ex. 3 0.22 Compound A 0.44 Meropenem 0.72 The above results indicate that the compounds of the present invention have strong antibacterial activity in the in vitro test and also have excellent effects in the treatment of infections in the in vivo test. Described specifically, compared with the aminomethyl compounds [compounds represented by the formula of the present invention wherein R1, R 2 and R 3 represent hydrogen atoms; Compounds A and B disclosed in Japanese Patent Application Kokai Hei 5-310740], the compounds of the present invention the compound of Example 3) exhibited superior activity against Pseudomonas aeruginosa. In addition, the compounds of the present invention exhibited excellent antibacterial activity against Pseudomonas aeruginosa 3719 which is resistant to Meropenem.

Furthermore, the compounds of the present invention the compound of Example 3) exhibited excellent pharmacokinetic properties such as half-life in blood.

When the compounds of the present invention were administered to rabbits, the incorporation in to the renal cortex is relatively low. When the compound of Example 3 was intravenously administered to rabbits at a dose of 200 mg/kg, the rabbits did not exhibit nephrotoxicity.

97162p2.doc P79569/FP-9716(PCT)/tsa-ig/English translation/0 7 10.98 P:\OPER\PDB\24076-97.286 6/8/99 103 (Formulation Example 1) Injections In 5 ml of distilled water for injection, 500 mg of the compound of Example 3 were dissolved. The solution was allowed to pass through a filter for sterilizing the solution, followed by lyophilization to afford a lyophilized preparation for injection.

(Formulation Example 2) Tablets Compound of Example 3 50 mg Lactose 126 mg Corn starch 23 mg Magnesium stearate 1 mg 200 mg The above ingredients, each in the powdery form, were mixed, subjected to wet granulation with corn starch, dried and then tableted by a tableting machine, whereby tablets, each 200 mg, were prepared. The tablets so obtained can be coated with sugar, if necessary.

Throughout this specification and the claims which follow, unless the context requires otherwise, the word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps.

P:\OPER\PDB\24076-97.286 6/8/99 104 THE CLAIMS DEFINING THE INVENTION ARE AS FOLLOWS: 1. (1R,5S,6S)-6-[(1R)-1-hydroxyethyl]-1-methyl-2-[(2S,4S)-2-[(3R)-3methylaminomethylpyrrolidin-l-ylcarbonyl]pyrolidin-4-ylthio]-l-carbapen-2-em-3carboxylic acid or a pharmacologically acceptable salt or derivative thereof.

2. A composition for the prevention or treatment of bacterial infections, which comprises a pharmacologically acceptable diluent or carrier in combination with an effective amount of a 1-methylcarbapenem compound in accordance with claim 1 or a pharmacologically acceptable salt or derivative thereof.

3. Use of a 1-methylcarbapenem compound in accordance with claim 1 or a pharmacologically acceptable salt or derivative thereof for preparing a medicament for the prevention or treatment of bacterial infections.

4. A method for the prevention or treatment of bacterial infections, which comprises administering to a warm-blooded animal a pharmacologically effective amount of a 1-methylcarbapenem compound in accordance with claim 1 or a *o*o pharmacologically acceptable salt or derivative thereof.

DATED this 6th day of AUGUST, 1999 Sankyo Company, Limited DAVIES COLLISON CAVE Patent Attorneys for the Applicant o